Standard Operating Procedures Version: 1research.ntu.edu.sg/guides/Documents/Ethics/NTU ARF SOP Manual_V… · Standard Operating Procedures . Version: 1.1 . ... NTU-ARF-SOP-6: SOP

Animal Research Facility Standard Operating Procedures

Version: 1.1

Approved By: Assoc. Prof Tan Nguan Soon, Andrew

Approved Date: 22 July 2015

Review Date: 22 July 2018

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TABLE OF CONTENTS

3 Introduction 3 General Guidelines for Users and Visitors to NTU 3 Overview 3 Authorized Access 4 Entry 4 House Rules and Safety 4 Animal Ethics and Care 5 Charges 5 Documentation 5 Use of Procedure Rooms and Equipment 6 Animal Euthanasia 6 Waste and Carcass Disposal 7 Reporting Animal Welfare Concerns 7 Contact Information 7 General Animal Care and Use 8 Policies and Programs 8 Animal Care Policy 9 Animal and Cage Identification System 10 Contamination Control Policies 10 Occupational Health and Safety Program 11 Pest Control 11 Institutional Animal Care and Use Committee (IACUC) 13 Standard Operating Procedures 14 NTU-ARF-SOP-1: SOP for Entry and Exit of Animal Research Facility 19 NTU-ARF-SOP-2: SOP for Daily Care of Animals and Monitoring of Animal Holding and

Breeding Environment 22 NTU-ARF-SOP-3: SOP for Animal Quarantine and Acclimatization 24 NTU-ARF-SOP-4: SOP for Cage Changing for Rodent Cages 27 NTU-ARF-SOP-5: SOP for Euthanasia of Rodents 30 NTU-ARF-SOP-6: SOP for Rodent Anesthesia by Isoflurane or Injectable Drugs 35 NTU-ARF-SOP-7: SOP for the Dispense of Controlled Drugs 37 Standard Operating Procedures for Animal Ethics 38 NTU-ARF-SOP-8: SOP for Escaped Animals 40 Standard Operating Procedures for Animal Research Facility Maintenance 41 NTU-ARF-SOP-9: SOP for General Maintenance of Cleanliness at ARF 44 NTU-ARF-SOP-10: SOP for Change of Compressed Gas Cylinders 46 NTU-ARF-SOP-11: SOP for Receiving Bedding and Diet 49 NTU-ARF-SOP-12: SOP for Veterinary Care and Sick / Abnormal Animal Reporting 52 NTU-ARF-SOP-13: SOP for Health Monitoring of Animal Research Facility 63 NTU-ARF-SOP-14: SOP for the Request and Procurement of Animals from Local

Sources 67 NTU-ARF-SOP-15: SOP for Importing Animals 72 NTU-ARF-SOP-16: SOP for Exporting Animals 77 NTU-ARF-SOP-17: SOP for Animal Carcass Disposal 79 NTU-ARF-SOP-18: SOP for Animal Research Facility Waste Management 84 NTU-ARF-SOP-19: Emergency Response Plan of Animal Research Facility 89 NTU-ARF-SOP-20: SOP for Animal Bites, Scratches and other Animal-Related Injuries 92 NTU-ARF-SOP-21: SOP for Preparation of Acidified Drinking Water for Laboratory

Rodents

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Introduction The Nanyang Technological University (NTU) Animal Research Facility (ARF) Standard Operating Procedures (SOP) serve as a reference for the Institutional Animal Care and Use Committee (IACUC) members, ARF staff members and animal researchers who are conducting scientific research on animals at the NTU ARF. These SOPs covers the methods that adhere to the National Advisory Committee for Laboratory Animal Research (NACLAR), and are associated with the care and use of laboratory animals in ARF. The manual provides a reference to conduct procedures in a safe and responsible manner.

General Guidelines for Users and Visitors to NTU

OVERVIEW

The ARF is the only licensed building to hold and use animals for scientific research in NTU. It was established in 2002 by the School of Biological Sciences. ARF is a standalone 2-storeys building located directly opposite the School of Chemical and Biomedical Engineering (SCBE). It is connected via an underground tunnel to the School of Biological Sciences (SBS). Level 1 is a BSL1, specific pathogen-free facility with 7 small animal holding rooms and 4 procedure rooms. Level 2 is a conventional small animal facility with 3 animal holding rooms and a BSL2 laboratory. All animals on Level 1 are housed in individually-ventilated cage (IVC) systems. The ARF holds a BioBeam gamma irradiation device and IVIS SpectrumCT Pre-clinical In Vivo Imaging System. ARF is monitored by 24 hr CCTV.

AUTHORIZED ACCESS

Entry to the ARF is only permitted to authorized personnel. Each authorization will last for a maximum of two years. To be eligible for access, the personnel must fulfil the following:

1. Personnel working with animals must have the Responsible Care and Use of Laboratory Animals (RCULA) certificate.

2. Personnel working with animals must have a protocol or part of the researchers in the protocol that is approved by the NTU’s Institutional Care and Use Committee, NTU IACUC.

3. Personnel must submit a security access application form to the Laboratory Animal Facility Manager (LAFM).

4. Personnel working with rodents must be tetanus-vaccinated and Hepatitis B-vaccinated if working with human samples untested with Hepatitis B.

For IACUC matters please refer to the IACUC Secretariat, [email protected].

Visitors must seek approval from the Laboratory Animal Facility Manager before entry. The ARF management reserves the right to decline entry without proper approval. All personnel who will be working in the facility must undergo the Safety Induction conducted by SBS and facility orientation by the ARF management before commencing work. Visitors are allowed entry only with accompaniment of authorized personnel at all times.

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mailto:[email protected]

ENTRY

There are 2 main entrances to the ARF. The entrance to Level 1 can be accessed from the drive in from the main road (60 Nanyang Drive) or from the SBS underground tunnel. The entrance to Level 2 can be accessed from the main road.

Personnel shall refer to the SOP for Entry and Exit of Animal Research Facility for step-by-step instructions on the proper entry to the ARF. Personal Protective Equipment (PPE) and covered shoes must be worn upon entering the facility. PPE are available on both levels of ARF.

Personnel are allowed to bring a small set of tools and reagents for their procedures. For large equipment/devices, prior approval must be sought. These shall be decontaminated and the steps are described in the abovementioned procedure. Personal belongings such as backpacks and lunch bags are not allowed into the facility. No children under the age of 12 are allowed.

HOUSE RULES AND SAFETY

All personnel shall complete the online SBS Safety Induction for Lab Users course before commencing laboratory work in the ARF. Please refer to the “In-house Safety Rules for SBS Labs” and adhere to the rules described at all times.

All personnel shall practice good laboratory practices for the safety of the humans and animals. All tools must be labelled with the PI’s name. Reagents must bear the Globally Harmonized System Classification and Labelling of Chemicals (GHS), and include the PI’s name, date of generation and content name. Tools and reagents should always be removed from the ARF after every trip as it is a shared facility. However, the personnel may leave it in the procedure room for the day at their own risk if they are returning to continue their work the next day. The ARF does not bear the responsibility for lost items.

Level 1: Personnel shall enter and exit via the locker room if they wish to return to continue working in Level 1 on the same day. Carcasses can be discarded in the allocated refrigerator in the clean area. Personnel who have entered the dirty zone are not allowed back into the clean zone for the day. However, they may continue working in Level 2.

Level 2: Personnel shall enter and exit via the main entrance. Entry to Level 2 from the Level 1 lift lobby shall be minimized. However, exceptions can be made to personnel who require the lift for larger equipment such as liquid nitrogen Dewar.

Level 2 Fish Lab: Personnel from the fish lab group may enter Level 2 of the facility in gloves only. However, they are not permitted to other rooms other than the fish lab without PPE.

PPE should be worn at both Levels 1 and 2 lift area. No cardboard items are allowed in Level 1 of ARF.

ANIMAL ETHICS AND CARE

All animals used for scientific purposes must be done according to the National Advisory Committee for Laboratory Animal Research (NACLAR) Guidelines. The ARF provides provisions that are basic essentials for the animals. These include sterile cages with corn

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cob bedding, nestlets, autoclaved water and irradiated diet. All cages and animals are observed once daily by the animal caretakers. Each cage is changed once every 10-14 days, or as and when needed. Diet and water are provided ad libitum unless otherwise stated by the researcher. All animal holding rooms follows a 12 hour light-dark cycle.

All researchers are responsible for the wellbeing of their animals. All protocols must adhere to the NTU IACUC Policy. All cages must be labelled with the IACUC protocol number, PI’s name, strain, gender, birth date and other valid information on the cage cards provided.

CHARGES

Please refer to the Animal Care Charges document for more information:

http://research.ntu.edu.sg/guides/Documents/Ethics/NTU%20ARF%20Animal%20Charges%202014.pdf

DOCUMENTATION

Researchers shall monitor and document the number of animals approved for their use, breeding, weaning and euthanize records. These records are subjected to checks during the Agri-Food and Veterinary Authority of Singapore (AVA) during the annual inspection.

Prior approval must be given for every ingoing and outgoing animal. Please refer to the forms under “Ethics” in the following link for the relevant forms:

http://research.ntu.edu.sg/guides/Pages/FormsandDocuments.aspx

USE OF PROCEDURE ROOMS AND EQUIPMENT

All personnel shall keep the shared procedure rooms neat and tidy to create a safe working environment for all users. Personnel who fail to do so may be banned from the facility, especially if it compromises the safety of other personnel and/or animals. BSL1 work can be conducted on the bench top while work involving risk group 2 agents, i.e. BSL2 work shall be conducted in Biological Safety Cabinets. Provisions such as L-fold paper, 70% ethanol, trash bags, PPE are provided in all rooms.

The procedure rooms and equipment in ARF can be booked online via the Central Equipment Booking System - Facility Online Manager (FOM©):

https://cebs.ntu.edu.sg/fom/welcome

Personnel shall email the Laboratory Animal Facility Manager to request for booking access. Personnel shall use the rooms and equipment only within the reserved time.

The equipment which aid in the operations of the facility includes:

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http://research.ntu.edu.sg/guides/Pages/FormsandDocuments.aspx

https://cebs.ntu.edu.sg/fom/welcome

1. Autoclave 2. Fume Hood 3. Class II Biological Safety Cabinet (BSC) 4. Centrifuge 5. Cage Changing Station 6. Laminar Flow Hood 7. Rack Washer 8. Bottle Washer 9. BioBeam 8000 10. IVIS SpectrumCT Preclinical In Vivo Imaging System 11. Isoflurane Vaporiser / Aneesthetic Machine 12. Bedding Disposal Station 13. Carbon Dioxide Chamber

Please refer to SBS’s Safe Practices & Safe Work Procedures (SWP) for the operation of autoclave, fume hood, BSC and centrifuge.

All equipment is serviced at least once a year. All users must inform the lab staff if the equipment is faulty.

To use the IVIS SpectrumCT Preclinical In Vivo Imaging System, the user must be R1 licensed and must seek endorsement from the L5 license holder (Prof Klaus Erik Karjalainen, [email protected]). More information can be found here:

http://www.nea.gov.sg/services-forms/licences-permits-building-plan-clearances/radiation

ANIMAL EUTHANASIA

Depending on the protocol, the animals may be euthanized using different methods that are recommended in the NACLAR Guidelines. Carbon dioxide is provided in Procedure Room 2 on Level 1 and the BSL-2 laboratory on Level 2.

WASTE AND CARCASS DISPOSAL

Waste shall be discarded properly in the appropriate receptacle. Biohazard waste, including blood stained tissues, must be discarded as biohazard waste. Glass and sharps must be discarded in the sharps bin located in all procedure rooms. General waste bins are also provided at various locations for other non-biohazard waste such as torn gloves, tissue used for cleaning the table etc.

Researchers are responsible for liquid wastes. Liquid wastes, whether chemical or biohazard, must be contained in a carboy with secondary containment. Proper GHS labels should be used and include information such as PI’s name and mixture. Researchers must arrange disposal of liquid waste when about 80% full.

Carcasses must be disposed in a separate biohazard bag. Biohazard bags with carcasses shall be double-bagged and sealed with autoclave tape, then stored in the freezer located in the dirty area of Level 1.

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http://www.nea.gov.sg/services-forms/licences-permits-building-plan-clearances/radiation

REPORTING ANIMAL WELFARE CONCERNS If there are concerns related to issues with the care and use of laboratory animals for scientific research or animal husbandry, persons may contact the NTU IACUC ([email protected]). Confidentiality of informant will be protected.

CONTACT INFORMATION The address of the facility and contact information can be found below:

Address: Animal Research Facility

60 Nanyang Drive

Singapore 637551

In-charge: Ms Lee Shok Li

Designation: Assistant Manager, Animal Research Facility

(Previous title: Laboratory Animal Facility Manager)

Email: [email protected]

Office No.: 6316 2921

ARF Level 1: 6316 2944

ARF Level 2: 6316 2945

Emergency No.: 6790 4777

General Animal Care and Use

Under the Animal & Birds (Care and Use of Animals for Scientific Purposes) Rules, any research facility which uses animals for scientific reasons must be AVA-licensed. The ARF is compliant with the National Advisory Committee for Laboratory Animal Research (NACLAR) Guidelines on the Care and Use of Animals for Scientific Purposes. The ARF renews its license once every year in July after an annual inspection conducted by the Agri-Food and Veterinary Authority of Singapore (AVA).

For more information about the NACLAR Guideline, please visit the following website:

http://www.ava.gov.sg/AnimalsPetSector/CareAndUseAnimalsForScientificPurp/

Flow of command of legislation for animal research in Singapore:

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http://www.ava.gov.sg/AnimalsPetSector/CareAndUseAnimalsForScientificPurp/

Policies and Programs

Animal Care Policy As a general guide, the following points shall be adhered to at all times when using animals for scientific purposes at the ARF:

1. Compliance to the NACLAR Guidelines. 2. Avoidance of sudden or loud noise in the facility. 3. Minimal / avoidance of discomfort, distress and pain to the animals. 4. Use of appropriate sedation, analgesia and anesthesia. 5. Provision of sufficient veterinary care.

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6. Provision of diet and water shall be ad libitum, unless otherwise specified and justified in the approved protocol.

7. Provision of sterile cages with bedding and enrichment. 8. Adherence to the approved animal use protocol at all times.

Animal and Cage Identification System The NTU ARF allows the researchers to identify the animals using their own system. Usually ear tags or ear punches are the main choices of identification. However, all animals housed at the NTU ARF must be identified. To manage this, the NTU ARF management utilizes a standard set of cage cards which shall be used by all researchers who houses their animals here. The following information must be included on the cage card:

(a) PI’s name (b) IACUC No. (c) Strain (d) No. of animals in the cage

There are 5 different types of cards that can be used. The recommended uses for each card are described below:

FRONT BACK RECOMMENDED USE

• Newly arrived animals. • Any other mice weaned and

waiting to be used. • Animals in this cage will be

fed with NORMAL DIET.

• Breeding cages or cages with nursing mother and her pups.

• Cages with these cards will be fed with BREEDING DIET.

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• For animals used for experiments.

• Animals in this cage will be fed with NORMAL DIET.

• For animals that have experienced surgery.

• It is the RESEARCHER’S responsibility to check the animal’s well-being on the 1st, 2nd, 3rd, 10th and 15th day.

• Animals in this cage will be fed with NORMAL DIET.

• For sentinel rodents only. • Given normal diet. • Utilised by SBS ARF Staff

only.

Contamination Control Policies

All personnel must not have visited another animal research facility or be in contact with other rodents 3 days prior to entry at Level 1. The personnel must be quarantined for a period of 3 days before entering Level 1. Personnel who have gone to Level 2 for the day are not allowed back in Level 1 until the next day.

No cardboard boxes, with the exception of glove boxes, are allowed in Level 1 of the ARF. This is to prevent incoming pests and introduction of mold.

Occupational Health and Safety Program All personnel who have to handle animals must be Tetanus-vaccinated and personnel who handle human specimens must be Hepatitis B-vaccinated. All SBS ARF staff members are fully vaccinated.

Each protocol requires the PI to conduct risk assessments and declare any hazardous agents and materials that may potentially cause harm to animals and/or humans. PIs possess Safety Data Sheets of all materials used.

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Pest Control The NTU ARF engages an external pest control company to manage the pests of the building. A preventive and treatment program is employed by the company to help control wild rodents, cockroaches, common ants and mosquitoes.

Institutional Animal Care and Use Committee (IACUC)

The NTU’s Institutional Animal Care and Use Committee (IACUC) is responsible for the oversight the institution’s animal care and use program to safeguard laboratory animal welfare. Under the AVA’s Animal and Birds Act (Chapter 7), Animals and Birds (Care and Use of Animals for Scientific Purposes), NTU must comply with the National Advisory Committee for Laboratory Animal Research (NACLAR) Guidelines on the Care and Use of Animals for Scientific Purposes. All projects involving the use of animals must seek approval from the IACUC before starting the project. The IACUC includes animal scientists, non-scientists, a veterinarian and members from the public community not affiliated to NTU. The IACUC:

1. Evaluates and approve animal research protocols. 2. Conducts audits: (i) semi-annual animal program review, (ii) annual facility review. 3. Reports to the CEO/IO and make recommendations relating to animal welfare. 4. Attends to concerns of animal welfare compromise. 5. Trains staff.

Principal Investigators must submit the Animal Use Protocol (AUP) form to the IACUC Secretariat ([email protected]) for a pre-review. A summary of the approval process is shown below:

The IACUC meets once every 2 months to review the protocols submitted. AUP forms that are submitted less than 2 weeks before the meeting shall not be reviewed until the meeting after next.

The following document outlines the IACUC’s policy on several animal procedures.

http://research.ntu.edu.sg/guides/Documents/Ethics/IACUC%20POLICY_2014_V01.pdf

For any concerns with regards to animal welfare, please contact the IACUC at [email protected], or Tel: 6316 2921. Confidentiality of the informant will be protected.

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http://research.ntu.edu.sg/guides/Documents/Ethics/IACUC%20POLICY_2014_V01.pdf


Standard Operating Procedures

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Ref. No.: NTU-ARF-SOP-1 Version: 1 Date of Issue: 18-Feb-15 Next Review Date: 18-Feb-18

Title: SOP for Entry and Exit of Animal Research Facility

Audience: All staff members, users, visitors and vendors.

1.1 SCOPE AND PURPOSE

1.1.1 This document describes the procedure to enter and exit the ARF, including the proper steps to don and doff PPE.

1.1.2 The purpose of this SOP is to minimise the risk of the spread of pathogens that can spread from or to the external environment by all personnel.

1.2 ASSOCIATED DOCUMENTS/FORMS

1.2.1 N.A.

1.3 GENERAL INFORMATION

1.3.1 Access to the ARF is restricted to authorized persons only. All personnel who wish to work in the ARF must first obtain access from the Laboratory Animal Facility Manager.

1.3.2 Personnel who will be working with animals in the ARF must be RCULA-certified.

1.3.3 The address of the ARF described in this procedure is

Animal Research Facility 60 Nanyang Drive Singapore 637551

1.3.4 Covered shoes must be worn at all times in the facility.

1.3.5 All visitors/personnel are advised to not bring valuables to the facility. The ARF shall not be responsible for any lost items.

1.3.6 PPE that has been contaminated or torn shall be discarded and replaced with a new piece.

1.3.7 All personnel are required to record their names, PI/organisation, date, entry and exit times for each access.

1.3.8 No cardboard materials are allowed into Level 1.

1.4 DEFINITIONS

1.4.1 SPF: Specific-Pathogen-Free

1.4.2 PPE: Personal protective equipment

1.4.3 RCULA Certification: Responsible Care and Use of Laboratory Animals

1.5 RESPONSIBILITIES

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1.5.1 Laboratory Animal Facility Manager

1.5.2 Animal Caretakers

1.5.3 Researchers

1.5.4 Visitors

1.6 PROCEDURE

1.6.1 ARF Level 1 Entry Procedure

1.6.1.1 Before entering ARF Level 1, the personnel shall sign into the access log book provided at the entrance, stating the name, PI / organisation, time in and date.

1.6.1.2 At the entrance (SBS-01-AH12), the personnel shall step onto the sticky mat with both shoes flat on it, and take a few steps on the mat before stepping forward into the locker room. This shall remove most dust particles from the shoes.

1.6.1.3 The personnel can place their personal belongings into one of the provided lockers in the area. The personnel shall then keep the keys with him/her at all times and collect the belongings later.

1.6.1.4 The personnel shall follow the visual instructions on the wall to don on the PPE.

1.6.1.5 The PPE required are: isolation gown, latex/nitrile gloves, bouffant caps, anti-slip shoe covers and face mask.

1.6.1.6 The personnel shall put on the PPE in the following sequence: isolation gown, bouffant cap, and face mask then gloves.

1.6.1.7 If the personnel have items to bring into the facility, these items shall be transferred into the provided boxes in the locker room. Each item shall be decontaminated with 70% ethanol before placing them into the box. The box shall then be decontaminated in the same manner. The transfer box/basket can be left in a corner of the locker room.

1.6.1.8 The personnel shall then bring a pair of shoe covers to the barrier. With the back facing the entrance to the door, the personnel shall sit on the barrier and put on a shoe cover to the left shoe, disinfect the sole of the shoe cover with the 70% ethanol, then cross over the barrier with the left leg first. The same shall be done for the right leg. Stepping on the barrier is not allowed.

1.6.1.9 After crossing over with both legs, disinfect both gloves with 70% ethanol before entering the premise.

1.6.2 ARF Level 1 Exit Procedure (To Return to ARF in the same day)

1.6.2.1 If the personnel wish to return to continue their work for the day, he/she shall exit through the entrance.

1.6.2.2 Personnel must not have entered the dirty area. If the personnel have already entered the dirty area, he/she shall not return to the Level 1 ARF for the day.

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1.6.2.3 The personnel shall sit on the barrier and cross over to the locker room one leg at a time. Stepping on the barrier is not allowed.

1.6.2.4 Transfer the items back into the visitor’s/researcher’s receptacle if any. The personnel shall decontaminate the box before returning it to the original location.

1.6.2.5 The personnel shall remove the PPE and discard each piece directly into the trash bin located in the locker room. To minimize the generation of aerosols and contaminating dust particles, the PPE shall be removed in the following sequence, starting from the dirtiest PPE: shoe covers, isolation gown, bouffant cap, gloves, face mask.

1.6.2.6 To minimize the generation of aerosols/dust, the gloves shall be removed in the following manner:

Figure 1: Source: https://www2.worksafebc.com/i/PPE/PDFs/InfoSheets/PPE09-015.pdf

1.6.2.7 The personnel shall collect any belongings from the locker and is now ready to exit the locker room.

1.6.2.8 The personnel shall wash their hands thoroughly with soap and water upon exiting the locker room.

1.6.2.9 The personnel shall sign out in the access log book, indicating the time of the exit.

1.6.3 ARF Level 1 Exit Procedure (No return for the day)

1.6.3.1 Personnel who have completed work in Level 1 for the day can choose to exit through the dirty corridor. This is encouraged if the personnel have big loads of carcass, or dirty cages to transport to the dirty area for disposal.

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1.6.3.2 The personnel shall enter the dirty corridor via their respective animal holding room’s exit.

1.6.3.3 The personnel shall enter the dirty area through the door at the end of the dirty corridor.

1.6.3.4 The personnel shall proceed to the lift area to remove the PPE. The personnel shall follow the steps stated on 1.6.2.5 and 1.6.2.6.

1.6.3.5 The personnel are now ready to exit the dirty area and will not be allowed to return to Level 1 for the day.

1.6.3.6 The personnel shall return to the Level 1 main entrance to sign out in the access log book, indicating the time of the exit.

1.6.4 ARF Level 2 Entry Procedure

1.6.4.1 There are 2 entrances to Level 2: first is from the road and second from the lift lobby that connects from Level 1. Personnel who have completed work in Level 1 can proceed to Level 2 via the lift.

1.6.4.2 However, if the personnel are entering Level 2 from the street, the personnel shall first sign into Level 2 on the access log book, indicating the name, PI / organisation, and entry time and date.

1.6.4.3 The personnel shall don on PPE at the entrance area in the following sequence: isolation gown, bouffant cap, and face mask then gloves.

1.6.4.4 Personnel are allowed to bring in items that will be used for their work in their own receptacle.

1.6.4.5 Personnel who will be working in the fish laboratory shall put on gloves but are not required to don on other PPE. These personnel are not allowed in other animal holding rooms. Personnel who are on full PPE and working in other rooms are also not allowed in the fish laboratory.

1.6.4.6 The personnel can now enter the animal room corridor.

1.6.5 ARF BSL2 Laboratory Entry and Exit Procedure

1.6.5.1 Personnel shall enter BSL2 only when working with risk level 2 agents.

1.6.5.2 The personnel shall find PPE such as safety goggles and gloves and shoe covers in the changing area.

1.6.5.3 Already donning full PPE, the personnel shall put on safety goggles, another layer of shoe covers and gloves before entering the BSL2 laboratory.

1.6.5.4 To exit, the personnel shall proceed to the changing area to remove the outer layer of PPE in the following sequence: shoe covers, gloves, safety goggles.

1.6.5.5 The personnel are now ready to exit BSL2 laboratory and into the general Level 2 animal laboratory area.

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1.6.6 ARF Level 2 Exit Procedure

1.6.6.1 The personnel shall proceed to the main entrance to remove PPE. The personnel shall follow the steps stated on 1.6.2.5 and 1.6.2.6.

1.6.6.2 The personnel shall wash their hands thoroughly with soap and water before exiting the facility.

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Title: SOP for Daily Care of Animals and Monitoring of Animal Holding and Breeding Environment

Audience: Laboratory Animal Facility Manager, Animal Caretakers


2.1.1 This document describes the procedures for the daily care of the animals housed at the ARF. It also describes the procedure for monitoring the animal holding and breeding rooms.

2.1.2 The purpose is to ensure that the well-being of the animals is properly taken care of by the animal caretakers.


2.2.1 Room Monitoring Checklist

2.2.2 Vet Check Form


2.3.1 The animals housed in the Animal Research Facility are checked once daily, including weekends and public holidays.

2.3.2 The animal holding and breeding room conditions are checked and recorded in the same checklist.

2.3.3 For any procedure that requires the cage to be opened, it must be done in the cage changing station in the respective animal holding rooms.

2.3.4 Weekend and public holiday rosters shall be scheduled by the senior animal caretaker. At least 2 technicians shall be assigned to perform routine checks according to this SOP on those days.

2.4 DEFINITIONS

2.4.1 N.A.



2.5.2 Animal Caretakers/technicians

2.6 PROCEDURE

2.6.1 The technicians shall observe all animals daily for clinical signs of illness, injury or abnormal behaviour, including weekends and public holidays.

2.6.2 The technicians shall observe each cage and check for the following:

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i Adequate feed and water

ii Cage cleanliness

iii Animal well-being

iv Dead animals

v Overcrowding

2.6.3 If the cage is found to have inadequate food and water, the technician shall top-up the appropriate diet and water.

2.6.4 The technician shall do a cage change if the cage is too dirty to last until the next scheduled cage change.

2.6.5 If an animal is found sick, injured or exhibiting abnormal behaviour, the technician shall record this in the Vet Check Form and hand it to the facility manager. The attending veterinarian and PI of the animals shall be informed and they shall decide the fate of the animal.

2.6.6 If an animal is found dead, the technician shall pick up the carcass and place it in a biohazard bag. This bag should be labelled with the date found dead, protocol number and PI’s name. The technician shall place this bag in the refrigerator designated for carcass only.

2.6.7 The technician shall record the room temperature, room humidity, IVC System’s temperature and humidity onto the Room Monitoring Checklist.

2.6.8 All rooms shall be swept and mopped once a week with disinfectant. The racks and doors shall be wiped with disinfectant once a month.

2.6.9 If the room is due for sweeping, mopping, rack cleaning, these should also be indicated on the Room Monitoring Checklist.

2.6.10 Any abnormal observations shall be recorded in the Room Monitoring Checklist and the technician shall inform the facility manager.

2.6.11 If checks are done on a weekend of public holiday, both technicians shall first check the rooms on Level 1, the SPF level, before checking Level 2.

2.7 APPENDICES

2.7.1 Appendix I: Room Monitoring Checklist

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Title: SOP for Animal Quarantine and Acclimatization

Audience: Laboratory Animal Facility Manager, Animal Caretakers, Attending veterinarian, Researchers / PIs


3.1.1 This document describes the procedures for quarantine and acclimatization of rodents admitted into the ARF.

3.1.2 The purpose of the document is to ensure that all animals that enter the facility do not carry pathogens that may spread diseases to the other animals.


3.2.1 Stock Card


3.3.1 Animals that come from overseas or non-approved sources have to be quarantined. Animals that are in quarantine are isolated from the rest of the facility for at least 1 week or 7 days until they appear to be free of infectious diseases or pathogens.

3.3.2 Animals acquired by the Animal Research Facility have to acclimatize for a period of 5 days to the environment before being used after quarantine or if it they originate from an approved local source.

3.3.3 If the Rodent(s) received are sick/unwell/injured, they will also be quarantined for inspection by the Attending veterinarian during his next visit.

3.3.4 The Attending veterinarian, on his next visit, will inspect the affected rodent(s) in the quarantine room and come up with a diagnosis and action plan, if any.

3.3.5 Rodent(s) due for acclimatization will be house normally in their respective holding rooms. However, they will not be used for any experiment or procedure until the acclimatization period of 5 days.

3.3.6 The Attending veterinarian will suggest rodent(s) for quarantine if required, such as rodent(s) coming from a local source which had a disease outbreak recently in their facility.

3.4 DEFINITIONS

3.4.1 N.A.



3.5.2 Attending veterinarian

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3.5.4 Principal Investigators/Researchers

3.6 PROCEDURE

3.6.1 The technicians shall receive the animals from the quarantine room of the Animal Research Facility at Level 1.

3.6.2 The technicians shall transfer the animals from each section of the crate to an autoclaved clean cage. The autoclaved corn cob, nestlet, diet and water bottle will be provided with the cage inside the quarantine room. Details of the animals shall be written on the Stock Card such that the PI/researcher can identify the animals later on.

3.6.3 The technicians shall transfer the animals to be quarantined to the Isolator and they are observed for a period of 7 days.

3.6.4 Rodent(s) originating from local sources other than approved vendors shall be placed directly into the holding rooms and acclimatized for 5 days.

3.6.5 The animal technicians shall transfer the quarantined animals that appear to be well to their respective animal holding rooms.

3.6.6 If a rodent is found to be sick/unwell/injured when received, they will be quarantined and inspected by the Attending veterinarian on his next visit.

3.6.7 PIs/Researchers shall locate the cages of the rodents from the cage cards and will be allowed to use the animals after the quarantine/acclimatization period.

3.7 APPENDICES

3.7.1 Appendix I: Animal Stock Card

Appendix I: Animal Stock Card

FRONT BACK

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Title: SOP for Cage Changing for Rodent Cages

Audience: Animal Caretakers


4.1.1 This document describes the procedure of changing soiled individually-ventilated cages to new autoclaved cages for the laboratory rodents.

4.1.2 The purpose is to protect the personnel and the environment from potential pathogens which may be carried by the research animals and vice versa.


4.2.1 N.A.


4.3.1 All rodents at SPF Level 1 are housed in IVCs while only mice are housed in IVCs at Level 2. Rats are housed in open-top cages at Level 2.

4.3.2 All IVCs must be opened using the cage changing stations in animal holding rooms.

4.3.3 All cages are changed every 10-14 days, or as often as required.

4.3.4 Autoclaved cages containing a layer of corn cob bedding, 2 pieces of nestlet material and covers with filters are prepared and placed at the clean areas.

4.3.5 Autoclaved water bottles with acidified water are also prepared and placed in the clean area.

4.3.6 Bags of irradiated diet are located in the feed storage room, as well as in the animal holding rooms. There are 2 types of diet: maintenance diet and breeding diet.

4.3.7 Transport of cages, water bottles and diet shall be done using steel trolleys that are available in the animal holding rooms or along the corridors.

4.3.8 Full PPE must be worn at all times while doing cage changing.

4.4 DEFINITIONS

4.4.1 SPF – Specific Pathogen-Free

4.4.2 IVCs – Individually-ventilated cages

4.4.3 LAFM – Laboratory Animal Facility Manager

4.4.4 ARF – Animal Research Facility

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4.6 MATERIALS

a. Autoclaved cages with corn cob, 2 nestlets, cage grill and covers with filters

b. Autoclaved water bottles containing acidified water

c. Autoclaved cage card holders

d. Irradiated maintenance or breeding diet

e. Long forceps – to transfer rodents form one cage to another

f. Scoop – for scooping dirty bedding to sentinel cage

g. Plastic container for soaking of forceps

h. 70% ethanol

i. Paper towels

j. Small brush with dustpan

k. Steel trolley

4.7 PROCEDURE

4.7.1 Cage Changing for IVCs at Levels 1

4.7.1.1 The animal caretaker shall switch on the Laminar Flow Hood in the animal holding room to allow the air flow to balance for about 10 minutes.

4.7.1.2 The animal caretaker shall count the number of cages to change in the room and proceed to gather that number of autoclaved cages and autoclaved water bottles into the room.

4.7.1.3 The animal caretaker shall sanitise the hood surface with 70% ethanol and tissue. The gloves shall also be sanitized with 70% ethanol between every cage.

4.7.1.4 70% ethanol shall be poured into a rectangular container until it is enough to soak the tip of the large forceps.

4.7.1.5 The animal caretaker shall proceed to the first rack to obtain a dirty cage. Place this cage into the sanitized surface of the hood and remove the water bottle. The cover shall be opened and placed next to the cage, with the internal surface facing upwards. The same shall be done for the clean cage.

4.7.1.6 The cage grill shall be lifted with one hand, and the other hand shall flick the forceps to ensure that it is not dripping wet with the 70% ethanol. Alternatively, dry the forcep with a tissue.

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4.7.1.7 The animal caretaker shall pick each rodent by the base of their tail using the forceps and transfer them carefully into the new cage. The diet from the soiled cage shall also be transferred to the new grill, and topped up with fresh maintenance diet or breeding diet. For rat cages, transfer the rats by holding them by the base of their tail with hands.

4.7.1.8 If dead animals are found, animal care taker shall follow the SOP for disposal of animal carcasses. If sick animals are found, this shall be recorded onto the vet check form and on the cage card itself.

4.7.1.9 The animal caretaker shall take another new cage, place it in the hood and open it.

4.7.1.10 Using the scoop, a handful of dirty bedding shall be scooped from the dirty cage into the new cage. The same shall be done for the diet. This cage shall be the cage for the sentinels.

4.7.1.11 When all the animals have been transferred, the new cage shall be covered, locked. The cage card shall be transferred to a new cage card holder. The new cage shall be given a new water bottle and placed back to the original slot on the rack.

4.7.1.12 The animal caretaker shall cover the dirty cage with its original water bottle and placed onto a steel trolley.

4.7.1.13 This shall be continued until all the cages in the room have been changed.

4.7.1.14 The animal caretaker shall push the trolley of dirty cages into the dirty area carefully, without stepping into the corridor.

4.7.1.15 The animal caretaker shall check that all cages are slotted into the racks securely.

4.7.1.16 Using the small brush and dustpan, the animal caretaker shall sweep the hood to remove all debris, followed by sanitization with 70% ethanol.

4.7.1.17 The room floor shall be swept. The animal care taker shall also update the daily log sheet to record that cage changing was done.

4.7.2 Cage Changing for IVCs at Level 2

4.7.2.1 This is similar to the steps described in 7.1.

4.7.2.2 However, cages with biohazard labels must be changed independently from the cages without biohazard labels.

4.7.2.3 The trolley of dirty cages with biohazard labels shall be placed onto a trolley with biohazard label. This trolley shall be pushed to the cage wash area when ready.

4.7.3 Cage Changing for Open-Top Cages at Level 2

4.7.3.1 As Level 2 is a conventional clean animal facility, cage changing does not have to be done in a laminar flow hood. The steps are similar to those described in 7.1 but done on an empty trolley instead.

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Title: SOP for Euthanasia of Rodents

Audience: Animal Caretakers, Animal Researchers


5.1.1 The scope of this document describes the different acceptable methods for humane euthanasia of rodents. It is intended for all personnel trained in euthanasia of rodents.


5.2.1 N.A.


5.3.1 Euthanasia means “good death” and refers to the intentionally ending a life with the purpose of relieving pain and suffering.

5.3.2 Euthanasia techniques should result in rapid loss of consciousness and quickly followed by cardiac or respiratory arrest with the final loss of brain function.

5.3.3 Euthanasia techniques should minimize distress and anxiety experienced by the animal before loss of consciousness.

5.3.4 The euthanasia techniques described in this document are approved by the NTU IACUC. They include but are not limited to the following:

i. Non-physical Methods

a) CO2 administration

b) Overdose of inhalant anesthetic

c) Overdose of injectable barbiturate

ii. Physical Methods

a) Cervical dislocation

b) Exsanguination

5.3.5 For decapitation and exsanguination, prior approval from the NTU IACUC must be sought and they shall be properly described and justified in the Animal Use Protocol.

5.3.6 All animal carcasses shall be disposed according to the Standard Operation Procedure for Animal Carcass Disposal.

5.4 DEFINITIONS

5.4.1 N.A.

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5.5.1 All personnel performing euthanasia on rodents

5.6 MATERIALS

i) Euthanasia chamber

ii) CO2 gas cylinder

iii) Isoflurane machine

iv) Isoflurane

v) Euthanasia solution

vi) Needles and syringes

vii) Decapitation device

viii) Sharps container

ix) Biohazard bag to contain animal carcass

5.7 PROCEDURE

5.7.1 CO2 Administration

5.7.1.1 The personnel shall ensure that the CO2 gas cylinder contains sufficient amount of CO2 to complete the euthanasia procedure.

5.7.1.2 The personnel shall place the animal(s) in the dedicated euthanasia chamber or cage. The chamber shall not be overcrowded, i.e. each animal has ample space to stand on the floor of the chamber/cage with all four feet, and is able to turn around with normal posture.

5.7.1.3 The personnel shall connect the compressed CO2 gas to the chamber via a hose and ensure that the chamber/cage is properly closed.

5.7.1.4 The tank valve shall be turned on and the CO2 shall induce rapid unconsciousness to the animals. The rodent shall immediately collapse to the floor of the chamber and begin to pant.

5.7.1.5 The gas flow shall sustain for a minimum of 1 minute until respirations have stopped.

5.7.1.6 The personnel shall ensure that the animal(s) is/are dead before removing them from the chamber. No respiratory movement shall be observed for at least 2 minutes.

5.7.1.7 If the animal is not dead, quickly cervical dislocate the animal.

5.7.2 Overdose of inhalant anesthetic

5.7.2.1 The animal(s) shall be placed into a euthanasia chamber/cage.

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5.7.2.2 The personnel shall connect the isoflurane machine to the chamber and ensure that the chamber/cage is properly closed.

5.7.2.3 The animal(s) shall be exposed to the vaporized isoflurane until respiration stops for at least 2 minutes.


5.7.3 Overdose of injectable barbiturate

5.7.3.1 Personnel can obtain Valabarb (sodium pentobarbitone 300mg/ml) from the Laboratory Animal Facility Manager.

5.7.3.2 120mg/kg of the solution is recommended for intraperitoneal (IP) injection.

5.7.3.3 The personnel shall scruff/secure the animal by hand or using an animal restrainer securely when injecting the euthanasia solution into the animal.

5.7.3.4 Respiration should cease for at least 2 minutes to ensure the death of the animal.


5.7.4 Cervical dislocation

5.7.4.1 The personnel shall hold the base of the rodent’s tail and place it on a surface, ideally the grill of the cage such that the rodent can grip onto it if it is still conscious. The rodent shall face away from the personnel.

5.7.4.2 The personnel shall place the thumb and index finger on the neck or base of the skull. Alternatively, use a rod to place at the base of the skull, perpendicular to the body.

5.7.4.3 The personnel shall quickly pull the base of the tail while pressing the rod or thumb/index finger on the neck, causing the separation of the cervical vertebra from the skull.

5.7.4.4 Crushing of the skull is normal.

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Title: SOP for Rodent Anesthesia by Isoflurane or Injectable Drugs

Audience: Laboratory Animal Facility Manager, Animal Researchers


6.9.1 This document describes the safe and proper procedure to sedate rodents with inhalant gas or injectable drugs as a reference for researchers.

6.9.2 The purpose is to ensure that rodent anesthesia is achieved fully for the required purpose, such as surgery, and to warrant the safe use of the isoflurane.


20.2.1 Isoflurane SDS

20.2.2 Ketamine SDS

20.2.3 Xylazine SDS


6.3.1 General Anesthesia is defined as a loss of consciousness coupled with loss of sensation and reflexes that is medically induced by one or more anesthetic agents, usually for a limited period of time.

6.3.2 There are 2 general anesthesia methods described in this SOP: anesthesia by inhalant isoflurane and injected Ketamine/Xylazine cocktail.

6.3.3 An anesthesia machine is located at each level of the ARF: SBS-01-AH13 and SBS-02-AH03. Isoflurane can be obtained from one of the animal caretakers. Ketamine and Xylazine can be obtained from the LAFM.

6.3.4 Ketamine and Xylazine are controlled drugs and their issuances are recorded by the LAFM.

6.3.5 Inhalation anesthesia by isoflurane for animals is administered with oxygen. It is both an anesthetic and analgesic.

6.3.6 Anesthesized rodents may lose body heat quickly. It is essential to provide a warmer / heating source until the animals recover from the anesthesia.

6.3.7 The animal must be fully sedated before the surgical procedure is performed. The onset of the sedation can be demonstrated by loss of pedal withdrawal reflex.

6.3.8 Analgesia should be administered before the surgical procedures as a pre-emptive measure. Alternatively, it can be provided immediately after the surgery is completed.

6.3.9 The induction chamber for isoflurane anesthesia should be cleaned before and after use, to avoid contamination.

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6.3.10 The researchers are responsible for post-surgical monitoring. Animal technicians shall contact the personnel with the emergency contact stated on the “EXPERIMENTAL” card. The “Surgical Checklist” card shall also be used for the researcher’s recording. The animals must be monitored for at least 3 days after the procedure for signs of pain, distress, eating and drinking.

6.3.11 The Attending Veterinarian shall refer to these cards during routine checks. He can be contacted as follows:

Name: Dr Frederic Chua

Email: [email protected]

Mobile: 9863 4258

6.3.12 All procedures must be recorded and include the details pertaining to the animal and protocol, as well as the following:

a. Weight

b. Agent used, dosage and route of administration

c. Time of anesthesia induction

d. Time of recovery from anesthesia

6.4 DEFINITIONS



6.4.3 SDS – Safety Data Sheet


6.5.1 Researchers

6.6 MATERIALS

6.9.1 Inhalant option:

6.6.1.1 Isoflurane solution

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6.6.1.2 Oxygen supply

6.6.1.3 Anesthetic Vaporizer

6.6.1.4 Induction Chamber

6.6.1.5 Nose cone of the appropriate size

6.6.1.6 Warmer / heat pad

6.9.2 Injectable option:

6.6.2.1 Ketamine and Xylazine solution

6.6.2.2 Water for injection (sterile water)

6.6.2.3 Syringes and needles of appropriate bore size

6.6.2.4 Warmer / heat pad

6.7 PROCEDURE

6.7.1 General Anesthesia by Isoflurane

6.7.1.1 The personnel shall turn on the oxygen valve and ensure that there is sufficient oxygen by checking the pressure gauge. If the pressure gauge is less than 200 PSI, contact one of the animal technicians to have it changed.

6.7.1.2 The personnel shall check the level of the isoflurane in the vaporizer. The ball should be at the half-way mark of the indicator for sufficient amount of isoflurane.

6.7.1.3 If the level of isoflurane falls below the halfway mark, the personnel shall refill isoflurane, ensuring that the vaporizer is in the off position. The personnel shall obtain a bottle of isoflurane from the animal technicians. Firstly, unscrew the cap of the anesthesia reservoir, remove stopper from the isoflurane bottle then pour carefully into the vaporizer reservoir until the ball reaches the half way mark. Replace the reservoir cap and place the stopper back onto the isoflurane bottle. The vaporizer is now ready for use.

6.7.1.4 The personnel shall pre-charge the induction chamber for about 5 minutes before placing the animal in it. Adjust the isoflurane vaporizer to 5% and oxygen to 0.8 – 1.2L/min for the pre-charge.

6.7.1.5 After 5 minutess, the personnel shall place the animal gently into the induction chamber and cover it properly. To induce anesthesia, the personnel shall adjust isoflurane vaporizer dial setting accordingly:

Mice: 1-2% of isoflurane, 0.8 – 1.2 L/min of oxygen

Rats: 2-3%, ~1.5 L/min of oxygen

The personnel shall allow about 5 mins for the induction. Subsequently, transfer the animal to a surgical bench and wear a nose cone over the

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mouth and nose of the animal, ensuring that the nostrils are not blocked by the nose cone.

6.7.1.6 The personnel shall check for the animal’s reflexes. The animal should not blink when the inner corner of the eye is gently touched, nor should it withdraw its feet when pinched. When blink reflex and pedal withdrawal reflex are both absent, surgery may commence.

6.7.1.7 The personnel shall assess the animal every 5 minutes during surgery. The breathing should be regular and the mucous membranes of the animal should remain pink.

6.7.1.8 After surgery, the personnel shall remove the nose cone and allow the animal to recover in the cage. Observe for any signs of distress before returning the cage to the animal holding room. Moistened food pellets can be placed on the bedding to encourage eating.

6.7.2 Anesthesia by Xylazine-Ketamine Injection

6.7.2.1 Weigh the animal and calculate the dosage required based on the recommended dosages below.

Recommended dilutions of injectable anesthetic cocktail for use in mice and rats: Mice Dosage: Ketamine – 120 mg/kg Xylazine – 16 mg/kg Dose Rate: 0.1 ml per 10 g of body weight by IP injection Duration of Effect: ~20 – 40mins Needle: 2

Stock/solution Volumes: Ketamine at 100mg/ml 0.60 ml Xylazine at 20 mg/ml 0.40 ml

Water 4.00 ml Total volume 5.0 ml = 50 doses

Rats Dosage: Ketamine – 100 mg/kg Xylazine – 10 mg/kg Dose Rate: 0.2 ml per 100 g of body weight by IP injection Duration of Effect: ~20 – 50mins

Stock/solution Volumes: Ketamine at 100mg/ml 2.0 ml Xylazine at 20 mg/ml 1.0 ml

Water 1.0 ml Total volume 4.0 ml = 20 doses

The drug cocktail can be stored at 4°C for a maximum of 2 weeks.

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6.7.2.2 To administer the drugs via IP injection, the animal shall be restrained using the proper techniques. The animal’s head shall be tilted downwards, exposing the abdomen to the personnel. Typically, inject into the lower right quadrant as it is less likely to penetrate a major organ. For rats, one person should restrain while another inject.

Figure 2: IP injection of a mouse.

Figure 3: IP injection of a rat.

6.7.2.3 After confirming that the animal is fully sedated by testing the reflexes, the surgery may commence. The personnel shall continue to check for the vital signs of the animal.

6.7.2.4 After surgery, the animal can be returned to the cage. The personnel shall allow the animal to recover and observe for signs of pain and distress before returning the cage to the animal holding room. Moist pellets shall be placed on the bedding to encourage eating and drinking.

6.8 DOCUMENTATION

6.8.1 All associated documents to do with the export shall be kept a minimum of 3 years.

6.9 REFERENCES

6.9.1 Institutional Animal Care and Use Committee, 2014. IACUC Guidelines: Anesthesia [online]. Iowa: The University of Iowa. Available at: http://animal.research.uiowa.edu/iacuc-guidelines-anesthesia [Accessed 11 June 2015].

6.9.2 Cold Spring Harbor Protocols, 2015. Recipe: Ketamine/Xylazine [online]. New York: Cold Spring Harbor Laboratory Press. Available: http://cshprotocols.cshlp.org/content/2006/1/pdb.rec702.full?text_only=true [Accessed 11 June 2015].

6.9.3 Frederick National Laboratory for Cancer Research, 2015. Ketamine Combinations for Rodent Anesthesia [online]. Maryland: The NCI at Frederick. Available at: https://ncifrederick.cancer.gov/Lasp/Acuc/Bethesda/Media/Documents/KetamineCombinations.pdf [Accessed 11 June 2015].

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6.9.4 Washington College, 2012. Standard Operating Procedure No. 1 Anesthesia and Analgesia in Rodents [online]. Maryland: Washington College. Available at: https://www.washcoll.edu/live/files/1227-no-1-anesthesia-sop-revised-1012pdf [Accessed 11 June 2015].

6.9.5 The Pennsylvania State University, 2013. Injectable Anesthesia [online]. Pennsylvania: The Pennsylvania State University. Available at: http://www.research.psu.edu/arp/anesthesia/injectable-anesthesia [Accessed 11 June 2015].

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Title: SOP for the Dispense of Controlled Drugs

Audience: Laboratory Animal Facility Manager, Animal Researchers


7.1.1 This document describes the procedure for dispensing controlled drugs to the animal researchers and PIs.


7.2.1 Anaesthetic Drugs Records 2014 (book)


7.3.1 The drugs dispensed using this SOP are all controlled drugs.

7.3.2 The drugs issued to the researcher/PI must be already approved in the IACUC protocol.

7.3.3 These drugs are purchased from the Attending veterinarian.

7.4 DEFINITIONS

7.4.1 Controlled Drugs: Drugs whose use and distribution is firmly controlled by the government due to its abuse potential or risk.





7.6 PROCEDURE

7.6.1 When a controlled drug is low in stock (~ <10mL left) or in request, the facility manager shall request to purchase this drug from the attending veterinarian.

7.6.2 Details of the purchased controlled drug (such as Ketamine, Xylazine etc.) are recorded into the Anaesthetic Drugs Records 2014 according to Appendix I. The details include date of purchase, date received, drug name, volume and concentration, quantity, bottle ID and drug expiry date.

7.6.3 These drugs shall be kept in a locked cabinet. The facility manager shall lock the cabinet and keeps the key at all times.

7.6.4 When a controlled drug is requested by a researcher or PI, they shall inform the facility manager in advance before collection.

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7.6.5 The facility manager draws the requested amount from the bottle into either a microfuge tube or centrifuge tubes, using a syringe and needle.

7.6.6 The tube shall be labelled with the drug name and the expiry date.

7.6.7 Details of this request shall be recorded into the abovementioned book, which includes information such as amount left in the bottle, and amount drawn.

7.6.8 When the requestor collects the drug, he or she shall sign the book to declare the collection.

7.6.9 This book shall be regularly checked and acknowledged by the attending veterinarian.

7.7 PROTECTIVE EQUIPMENT

7.7.1 The facility manager who dispenses the drugs is advised to wear a pair of protective gloves when drawing and dispensing the drugs.

7.8 DOCUMENTATION

7.8.1 This Anaesthetic Drugs Records shall be kept a minimum of 3 years.

7.9 APPENDICES

7.9.1 Appendix I: List of Drugs in Stock (Template)

Appendix I: List of Drugs in Stock

List of Drugs in Stock

No. Ref No. Date In Supplier Bottle ID Brand Description Concentration Volume Expiry Date

Sign In

Date Started

Refer to page

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Animal Research Facility | School of Biological Sciences

Reg. No. 200604393R

Standard Operating Procedures for

Animal Ethics

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Reg. No. 200604393R

Ref. No.: NTU-ARF-SOP-8 Version: 1 Date of Issue: 8-Dec-14

Prepared by: Lee Shok Li Approved by: Dr Andrew Tan

Next Review Date: 8-Dec-17

Title: SOP for Escaped Animals

Audience: All animal handlers.


8.1.1 This document describes the procedure for management of animals that have escaped from their cages and animals of unknown origin are found within the NTU’s Animal Research Facility.

8.1.2 The purpose is to reduce the possibility of exposing research colonies to pathogens and diseases either from another colony or animals of unknown origin.


8.2.1 N.A.


8.3.1 IVCs are opened in cage changing stations in animal holding rooms. However, they are opened on bench tops in procedure rooms.

8.3.2 Room doors should always be closed before conducting animal work to reduce the chance of an escaped rodent to the common corridor.

8.3.3 Full PPE shall be worn when handling escaped rodents.

8.4 DEFINITIONS




8.4.4 PI – Principal Investigator





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Reg. No. 200604393R

8.5.3 Researchers

8.5.4 Attending Veterinarian

8.6 PROCEDURE

8.6.1 If an escaped rodent is found within an animal holding room or procedure room, the door shall be shut to prevent it from escaping into the corridor.

8.6.2 The personnel shall attempt to catch the rodent. As the rodent will be stressed and may bite or scratch in defense, an extra layer of gloves is recommended before attempting to catch it. Once caught, the animal must be housed in a new cage separately.

8.6.3 The LAFM and the PI shall be informed.

8.6.4 The PI and/or designee shall attempt to identify the rodent through the ear marks, location, coat colour and/or the absence of known research animals.

8.6.5 If the animal is identified, the cage which it escaped from shall be updated with the new number of animals, and the date of the escape.

8.6.6 The PI shall discuss with the Attending Veterinarian and the LAFM if certain conditions should be in place regarding the future use of this animal. For example, this animal may never be housed together with other animals until the end-point.

8.6.7 If the animal cannot be identified, it shall be euthanized by an Animal Caretaker.

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Reg. No. 200604393R

Standard Operating Procedures

for Animal Research Facility Maintenance

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Reg. No. 200604393R

Ref. No.: NTU-ARF-SOP-9 Version: 1 Date of Issue: 18-Feb-15

Prepared by: Lee Shok Li Approved by: Dr Andrew Tan

Next Review Date: 18-Feb-18

Title: SOP for General Maintenance of Cleanliness at ARF



9.1.1 This document describes the steps taken to maintain cleanliness and the health status of the animals at the ARF.

9.1.2 The purpose is to ensure that all animal caretakers do the same procedures to effectively maintain the ARF.


9.2.1 Room Check Sheet


9.3.1 All animals are checked at least once daily, including weekends and public holidays.

9.3.2 Weekend and public holiday duties are done on a rotational basis. The rosters are planned by one of the senior animal caretakers (Manis).

9.3.3 All cages must be handled in the cage changing station.

9.3.4 The animal caretakers should sanitise the gloves with 70% ethanol or hand sanitizer between cages. For cages suspected of a contamination, a change of gloves should be done before and after handling the cage.

9.3.5 Diets, corn cob and nestlets are stored at SBS-01-AH09 and SBS-02-AH07.

9.3.6 Detergents are stored in SBS-01-AH16 and SBS-02-AH02.

9.3.7 As of ??? , no cardboard boxes are allowed in ARF L1l, with the exception of glove and face mask boxes. These shall be discarded during routine housekeeping.

9.4 DEFINITIONS

9.4.1 N.A.



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Reg. No. 200604393R

9.6 PROCEDURE

9.6.1 Animal Room Maintenance

9.6.1.1 All animals shall be checked at least once daily, including weekends and public holidays.

9.6.1.2 Each cage shall be observed by visual inspection for sufficient diet pellets and water in the water bottles. If necessary, open the cage in the cage changing station to check for injured or distressed animals, new born pups or dead animals.

9.6.1.3 The animal caretaker shall replace the water bottle with a fresh one and top up the diet in the cage if either is found to be insufficient. All cages shall be given the maintenance diet unless it has a breeding cage card, or as specified by the researcher.

9.6.1.4 The bottles that are replaced shall be placed on a trolley for dirty cages. This trolley shall be pushed in to the dirty area when full.

9.6.1.5 The animal caretaker shall record any actions taken or observations in the daily log sheet. If a sick/injured/distressed animal was observed, the animal caretaker shall fill up a vet check form and hand it to the LAFM. In addition, the animal caretaker shall write the observation on a post it and paste it on the cage card to keep the researcher informed.

9.6.1.6 The room humidity and temperatures shall also be recorded by reading the room’s thermostat and hygrometer. The same shall be done for the IVC systems.

9.6.1.7 The animal caretaker shall clean and sanitse the cage changing station after each use.

9.6.2 Procedure Room Maintenance

9.6.2.1 The procedure benches and worktables shall be sanitized daily and/or after each use.

9.6.2.2 The animal caretakers and researchers shall keep the rooms neat and tidy as they are shared between users.

9.6.3 General Maintenance

9.6.3.1 All floors, including animal rooms, procedure rooms, and corridors and changing areas, shall be swept daily and mopped with detergent once a week, usually on a Friday.

9.6.3.2 All door handles shall be sanitized weekly.

9.6.3.3 All general waste bins shall be checked daily. When full, the trash bag shall be removed and replaced with a new trash bag. The full trash bag

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Reg. No. 200604393R

shall be tied and disposed into the garbage bin located at Level 1 unloading area.

9.6.3.4 All rooms, including animal holding and procedure rooms, shall be well-supplied with gloves, face masks, M-fold papers and 70% ethanol.

9.6.3.5 All animal holding rooms shall be well-supplied with the essential diets, cage cards, and vet check forms.

9.6.3.6 All procedure rooms shall be well-supplied with sanitizing wipes and biohazard bags.

9.6.3.7 The changing area shall be well-supplied with sanitizers, 70% ethanol, M-fold papers, face masks, bouffant caps, gloves, isolation gowns and non-slip shoe covers.

9.6.3.8 The sticky mat at the entrance of the L1 changing area shall be changed daily.

9.6.4 Logistics

9.6.4.1 All supplies should be available to last for at least another month. The animal caretakers shall inform the LAFM as soon as the supplies fall below the amount required for another month’s use.

9.6.4.2 All animal caretakers shall ensure that the supplies used are not expired, unless specified to be safe by the supplier. This must be justified with a letter from the manufacturer.

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Title: SOP for Change of Compressed Gas Cylinders

Audience: Animal Research Facility users, Animal Caretakers


10.1.1 This document describes the steps taken for the safe changing of compressed gas cylinders in the Animal Research Facility.

10.1.2 The purpose is to ensure that the users are aware of the risks involved and are familiar with handling of compressed gas cylinders.


10.2.1 N.A.


10.3.1 For more information, refer to the Guideline on Compressed Gas Cylinder Safety by the Office of Health and Safety, NTU:

http://intranet.ntu.edu.sg/ohs/Shared%20Documents/OHS%20DOCUMENTS/Guidelines/GUIDE%20-Compressed%20Gas%20Cylinder%20Safety%201.2%20final.pdf

10.3.2 There are two types of compressed gas cylinders used in the ARF: medical oxygen and carbon dioxide.

10.3.3 Medical oxygen comes in 2 sizes: 50L and 10L while carbon dioxide comes in 50L only.

10.3.4 The medical oxygen is used for isoflurane anesthesia while carbon dioxide is used for euthanasia and CO2 incubators.

10.3.5 8 of 50L gas cylinders are located at SBS-02-AH10.

10.3.6 There are 2 indicators attached to the 50L gas cylinder.

10.3.7 The user should be familiar with the properties of the compressed gas and understand the risks.

10.3.8 The user must be trained or have passed the Safe Handling of Compressed Gas Cylinders course located at NTULearn.

10.3.9 The user must wear safety shoes when transporting gas cylinders.

10.4 DEFINITIONS

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10.4.1 N.A.




10.6 PROCEDURE

10.6.1 An empty cylinder tank is indicated by the low pressure of the pressure gauge.

10.6.2 The user shall transport a new gas cylinder to its destination using a gas cylinder hand truck. Ensure that the cylinder is secured to the truck with a chain or a belt. The valve-protection cap should be left in place at all times until it is ready to be changed.

10.6.3 The user shall ensure that the cylinder valve of the cylinder to be changed is closed securely and all pressure is released from the connected system. Purge all gases and remove the regulator or connector slowly using a spanner. Replace the cylinder cap once the regulator is removed.

10.6.4 The user shall remove the “IN USE” section of the tag and this tank is now “EMPTY”. However, the animal caretaker shall treat all cylinders as if residual gas remains in the cylinder.

10.6.5 The user shall release the “EMPTY” tank from the safety chain, and then place the new tank into the gas bracket.

10.6.6 The user shall connect the new tank to the tube or connector securely using the spanner.

10.6.7 The user shall remove the “FULL” section of the tag of the new cylinder such that it now shows “IN USE”.

10.6.8 The “EMPTY” tank shall be placed onto the hand truck and secured with the chain or strap. In a safe manner, this shall be transported back to the gas cylinder storage area until it is ready for vendor’s collection.

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Title: SOP for Receiving Bedding and Diet



11.4.1 This document describes the procedures of receiving bedding and diet for the research animals in the Animal Research Facility.

11.4.2 The purpose is to ensure that the staff members are familiar with the correct posture of unloading heavy bags from the truck.


11.2.1 Bedding Movement Log

11.2.2 Diet Movement Log


12.3.1 The current supplier of bedding and diet is Biosystems Corporation (GeBIZ Contract Number: NTU000ECO13002903) and the contract is for a period of approximately 2 years.

12.3.2 The bedding used is Corn Cob Bedding; Granule Size: 5-7mm (1/4”); Bag Size: 18kg (40lb).

12.3.3 The normal diet used is Altromin #1324 Maintenance Diet for Rat/Mice; Irradiated; Bag Size: 10kg double layer plastic packing.

12.3.4 The breeding diet used is Altromin #1314 mod 9% fat Breeding Diet for Rat/Mice; Irradiated; Bag Size: 10kg double layer plastic packing.

11.4 DEFINITIONS

11.4.1 N.A.




11.6 PROCEDURE

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11.6.1 The technician in-charge of purchases shall arrange for the delivery time and date

of the bedding and/or diet.

11.6.2 The deliverer shall call the technicians before arrival.

11.6.3 Upon arrival, all technicians who are available shall proceed to the bedding store on Level 1 to assist with the unloading of the bags from the unloading bay.

11.6.4 The deliverers shall unload the bags from the truck, and pass them to the technicians who shall sanitise each bag with ethanol before arranging them on the shelves in the room.

11.6.5 Several bags are loaded onto a trolley by another technician and brought to level 2 via the lift at the second entrance. They shall be stored in the bedding store and diet store in Level 2.

11.6.6 The technicians shall use proper lifting techniques during unloading to avoid injuries.

Figure 4: http://www.yourdrgreens.com/blog/2011/04/are-you-aware-of-your-posture/

11.6.7 Upon receipt of the bags, the technician shall verify the number of bags and fill in the bedding and animal feed movement logs (Appendix I and Appendix II).

11.6.8 Details such as batch numbers and expiry dates shall be recorded.

11.7 DOCUMENTATION

12.7.1 All movement logs shall be kept for a minimum of 3 years.

11.8 APPENDICES

12.8.1 Appendix I: Animal Diet Movement Log

12.8.2 Appendix II: Bedding Movement Log

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Appendix I: Animal Diet Movement Log

Appendix II: Bedding Movement Log

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Title: SOP for Veterinary Care and Sick / Abnormal Animal Reporting

Audience: Animal Research Facility users, Animal Caretakers, Attending veterinarian


12.1.1 This document describes the procedures of reporting any sick/abnormal rodents as well as the follow-up actions if any. In addition, it describes the actions that the Attending veterinarian shall take in response to cases of sick or distressed animal.

12.1.2 The purpose of this SOP is to document all


12.2.1 Vet Check Form (ARF_VETCHECK01)


12.3.5 Rodent(s) that are found to be sick / injured or are exhibiting abnormal behavior shall be reported using the Vet Check Form.

12.3.6 The form shall be communicated between the person who made the discovery, the facility manager, the attending veterinarian and the PI/Researcher of the project involving the animal.

12.4 DEFINITIONS

12.4.1 Sick / abnormal rodents: Mice or rats that appear to have behavioural and physiological abnormalities.




12.5.3 Animal Care-takers


12.6 PROCEDURE

12.6.1 Rodent(s) inside the facility that are found to be abnormal (sick, injured, abnormal behavior, etc) must be documented using the Vet Check Form.

12.6.2 The Vet Check Form can be found on the basket trolley inside each animal holding room.

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12.6.3 The Vet Check Form must be duly filled before submission to the facility manager.

Additional details or observations made, if available, should be indicated in the remarks column of the form.

12.6.4 The semi-completed form shall be submitted to the attending veterinarian via the facility manager.

12.6.5 The attending veterinarian shall locate these rodents using the form, perform diagnosis and recommend any follow-up action plans onto the same form. This shall be handed back to the facility manager.

12.6.6 The facility manager shall email this form to the relevant PI/Researcher.

12.6.7 The PI/Researchers will follow the recommendations made by the attending veterinarian, if any.

12.7 DOCUMENTATION

12.7.2 All vet check forms shall be kept a minimum of 3 years.

12.8 APPENDICES

12.8.3 Appendix I: Vet Check Form

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Appendix I: Vet Check Form

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Title: SOP for Health Monitoring of Animal Research Facility

Audience: Animal Research Facility users, Animal Caretakers, Attending veterinarian


13.1.1 Animals that come from overseas or non-approved sources have to be quarantined.


13.2.1 Sentinel Cage Card

13.2.2 Approved IACUC Protocol for Sentinel Program

13.2.3 Pro-forma Invoice

13.2.4 USDA Statement

13.2.5 US Health Monitoring

13.2.6 Sample Collection


13.3.1 An approved IACUC protocol must be valid before animal sentinels can be purchased from approved vendors in Singapore. (IACUC Protocol No.: A0253)

13.3.2 Dirty-bedding sentinels are the selected method for the health monitoring of the rodents in the Animal Research Facility.

13.3.3 Biomed Diagnostics Singapore (local distributor for IDEXX BioResearch) is selected for the sentinel testing services from 2014 – 2015 (ITQ No.: NTU000ETQ14002069). Therefore, this SOP only applies to this vendor. It must be reviewed when a different vendor is selected or if the vendor changes the SOP for sample collection.

13.3.4 Sample kits are provided by Biomed Diagnostics Singapore.

13.3.5 Samples are all sent to IDEXX BioResearch (USA) for analysis.

13.3.6 ICR mice and Sprague Dawley® rats are the selected strains for this program due to their robust immune system. Only females are used.

13.3.7 The number of sentinels to order is dependent on the number of active sides of the IVC rack. There are 24 active racks for mice and 2 active racks for rats in the

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SPF level of the NTU Animal Research Facility. Therefore, there should be at least 24 mouse sentinels + 2 rat sentinels at any one time.

13.3.8 Each batch of sentinels is tested every quarter of the year. Refer to Appendix II, III and IV for the time line and panels tested.

13.3.9 Any transfer of mice between cages, collection of dirty bedding etc, shall be performed in the room’s cage changing station.

13.4 DEFINITIONS

13.4.1 Dirty-bedding Sentinels: Mice or rats used for detection of pathogens that may be present in the facility by exposing the animals to dirty bedding samples from other cages.





13.6 PROCEDURE

13.6.1 Maintenance of Dirty Bedding Sentinel Rodents

13.6.1.1 The facility manager shall purchase 24 ICR females + 2 Sprague Dawley® females from InVivos.

13.6.1.2 When the rodents arrive from the supplier in transport crates, the technician shall transfer each rodent to individual clean cages in the Level 1 quarantine room, using the cage changing station. Diet, nestlets and water are not necessary.

13.6.1.3 The technician shall immediately transfer any 4 mice to animal rooms 1, 3, 4, 5, 6 and 8 respectively and the 2 rats to room 7 using a trolley.

13.6.1.4 In each room, the technician shall transfer the new rodent into an existing sentinel cage, which already has an older sentinel in it.

13.6.1.5 During cage changing, the technician shall collect a handful of dirty bedding from every cage of the same side of the rack and pool into a clean empty cage. The same is done for the diet. The technician shall sanitise gloves with ethanol between cages at all times.

13.6.1.6 The cage with the collected dirty bedding and used diet shall be used as the sentinel cage. A sterile water bottle shall be provided to the sentinel cage. The sentinel cage change shall be done once every 10-14 days.

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13.6.1.7 The technician shall update the new sentinel’s information on the cage

card. It shall include the rodent’s ID, birthdate and gender.

13.6.1.8 The sentinel cage shall be placed back to the designated position of each respective rack. This position is usually the bottom right or left corner of the rack to standardize all sentinels.

13.6.1.9 A flow chart describing the maintenance of bedding sentinels is below:

13.6.2 Collection of Samples from Sentinel Rodents

13.6.2.1 The following materials are provided by the vendor:

i Opti-spot Strips

ii Wet culture swabs

iii Flocked swabs for mice

iv Flocked swabs for rats

13.6.2.2 The following materials are required for sample collection:

i Microfuge tube rack

ii Ziplock bags

iii Indelible Marker

iv Sterile microfuge tubes

v Sterile needles

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vi Virkon as disinfectant

vii Tissue/gauze

viii Clean forceps

ix Clean scissors

13.6.2.3 All sample collections are performed in the cage changing station located in each room.

13.6.2.4 Depending on the test panels that are scheduled, different types of samples have to be collected. The following tables summarise the samples required for each panel test, and the materials required for each type of sample collection.

Mouse HM Program

Month for HM IDEXX Panel Sample required Sep '14 Comprehensive Plus Blood Nov '14 Comprehensive Plus Blood Feb '15 Comprehensive Plus Blood

May '15 Global Blood Micro Panel 4 Wet oral swab, flocked oral swab, feces Mouse PCR Skin/Pelt swab, feces

Rat HM Program

Month for HM IDEXX Panel Sample required: Sep '14 Global Blood Nov '14 Global Blood Feb '15 Global Blood

May '15 Global Blood Micro Panel 4 Wet oral swab, flocked oral swab, feces Rat PCR Skin/Pelt swab, feces

Sample Materials Required for Sample Collection

Blood Opti-Spot strip, 25 gauge needle

Fur / skin swab Nylon swab (Regular tip), microfuge tube, scissors

Wet oral swab – culture Orange cap BBL culture swab with transport media

Flocked oral swab – PCR Flocked nylon swab (Mini tip), microfuge tube, scissors

Feces – culture, PCR, and parasitology microfuge tube, disinfected forceps

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13.6.2.5 The following flow charts describe the method for each sample collection.

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13.6.2.6 The different samples shall maintain in conditions according to the following table:

Sample Conditions

OptiSpot Blood Samples

• At RT and 4°C: 7 days • At -20°C: 1 month • Samples shall be sealed in plastic bag with a silica

desiccant pack at room temperature. Feaces and Culture Swabs

• Feces and culture swabs should be kept refrigerated for no longer than 24 hours prior to shipment.

• 2-8 °C (Cold shipment)

Fur / Skin Swab • Store in refrigerator for next day delivery. • For longer storage, freeze the sample in -20°C freezer

prior to shipment. • 2-8 °C (Cold shipment)

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13.6.2.7 The facility manager shall prepare the documents 2.3 – 2.6, sign, and

submit to the vendor. Pick up of samples shall be on the sample collection day itself or the next day.

13.6.2.8 The vendor shall analyse and generate the results within 7 working days. The results can be accessed via IDEXX’s online system.

13.7 DOCUMENTATION

13.7.1 All health monitoring records shall be kept for a 5 year period.

13.8 APPENDICES

13.8.1 Appendix I: Animal Stock Card

13.8.2 Appendix II: Mouse Sentinel Timeline

13.8.3 Appendix III: Rat Sentinel Timeline

13.8.4 Appendix IV: Test Panels

Appendix I: Animal Stock Card

FRONT BACK

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Appendix II: Mouse Sentinel Timeline

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Appendix III: Rat Sentinel Timeline

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Appendix IV: Test Panels

Mouse Microbiology Panel 4

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Rat Microbiology Panel 4

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Title: SOP for the Request and Procurement of Animals from Local Sources

Audience: Laboratory Animal Facility Manager, Animal Caretakers, Researchers / PIs


14.1.1 This document describes the procedure for purchasing and receiving rodents locally.


14.2.1 Animals Requisition Form (ARF_ANIMALREQUEST01)


14.3.1 This SOP applies to all rodent purchases and local animal transfers from other institutes.

14.3.2 The facility management reserves the right to reject the request.

14.3.3 The form above can be found in the NTU server.

14.4 DEFINITIONS

14.4.1 N.A.





14.6 PROCEDURE

14.6.1 When requesting or purchasing animals from local sources, the PI/researcher shall submit this form to the facility manager or assigned animal caretaker via email. The forms shall be duly filled, adhering to the terms and conditions.

14.6.2 The animal caretaker checks the form to ensure that there is enough space in the requested room to hold the animals requested.

14.6.3 The facility manager checks the form to ensure that the protocol stated on the form is still valid, and that the numbers requested do not exceed the number approved.

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14.6.4 The requestor will be notified via e-mail on the outcome of the approval. The

Animal Caretaker will keep a copy of the form, if approved.

14.6.5 The transfer or purchase of the rodents shall proceed upon approval.

14.6.6 The PI/researcher shall keep the facility manager and assigned animal caretaker informed of the actual arrival date.

14.6.7 Upon arrival, the animal caretaker shall transfer the mice to clean cages and transfer them to the respective animal holding room. He or she shall complete the form by verifying the animals received. This completed form shall be submitted to the facility manager along with the invoice and delivery order if any.

14.6.8 The facility manager shall keep the animal requisition form and hands the other documents to the relevant finance person of the institution, informing the PI/researcher at the same time.

14.7 DOCUMENTATION

14.7.1 All Animal Requisition forms shall be kept for a 3 year period.

14.8 APPENDICES

14.8.1 Appendix I: Animals Requisition Form

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Appendix I: Animals Requisition Form

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Title: SOP for Importing Animals

Audience: Laboratory Animal Facility Manager, Attending veterinarian, Researchers / PIs


15.1.1 This document describes the procedure for importing rodents from overseas sources into NTU’s Animal Research Facility.

15.1.2 The purpose is to ensure that the required documents and permits are properly obtained to ensure a smooth and legitimate import of animals.

15.1.3 This SOP also ensures that the animals that are imported do not carry pathogens that could potentially cause an infection to the animals at ARF.


15.2.1 Animals Import Form (ARF_ANIMALIMPORT01)

15.2.2 AVA Import Permit

15.2.3 AVA Veterinary Conditions for the Importation of Laboratory Animals (1/4) – Rabbits, Guinea-pigs, Hamsters, Mice, Rats, and Gerbils (1/2)

15.2.4 Animal Packing List


15.3.1 The abovementioned AVA documents can be found on the Agri-Food & Veterinary Authority of Singapore Website: www.ava.gov.sg.

15.3.2 The AVA Import Permit must be paid in full by the PI/researcher prior to the commencement of the import.

15.3.3 The PIs, facility managers and responsible veterinarians from both facilities must be informed of this import.

15.3.4 The courier that is selected by the PI/researcher to handle this import should advise the documents that are required on both sites for this export.

15.3.5 The AVA Import Permit is not transferable and is valid for only one consignment for up to 30 days from the date of issue. The AVA Import Permit and the original Veterinary Certificate have to accompany this import. Animals are only to be derived from the source as indicated in the AVA Import Permit.

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15.3.6 The animal packing list must be submitted to the facility manager prior to the

import.

15.3.7 The exporting facility’s latest relevant health check report must be submitted to the facility management prior as part of the approval process.

15.3.8 For further clarification with regards to the regulations required by governing authorities and animal import, seek the AVA and selected courier service for advice.

15.3.9 Rodents arriving from overseas will be quarantined in the Animal Research Facility’s Quarantine room for at least 1 week.

15.3.10 The facility manager and attending veterinarian reserves the right to reject any animals import.

15.4 DEFINITIONS

15.4.1 AVA: Agri-Food and Veterinary Authority of Singapore






15.6 PROCEDURE

15.6.1 The PI/researcher shall source and select a courier service to handle this import. Subsequently, this request shall be made by filling and submitting the Animals Import Form to the NTU’s facility manager ensuring that the terms and conditions listed on the form are adhered to. This is subjected to approval by the facility manager and attending veterinarian.

15.6.2 The PI/researcher shall apply and pay in full for the AVA Import Permit and the AVA Veterinary Certificate prior to the import. Once successfully applied, these shall be submitted to the facility manager for records.

15.6.3 The PI/researcher shall request the exporting facility to submit the latest relevant health check report to NTU’s facility management. This is reviewed by the attending veterinarian and subjected to approval.

15.6.4 In the event that the attending veterinarian decides to approve the import but on the condition that further animal testing is done when the animal reaches the animal research facility, the cost of these test(s) shall be borne by the PI/researcher.

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15.6.5 The facility manager and attending veterinarian shall review the request. The

outcome of the request will be informed by email.

15.6.6 The animal packing list shall be submitted to the facility manager, who assigns it to one of the animal caretakers.

15.6.7 Upon arrival, the animal caretaker receives the animals and checks them with the animal packing list. The animals shall be transferred to sterile cages in accordance with the Quarantine SOP. They will be quarantined for a week.

15.6.8 If further animal testing is required, the animals shall be quarantined until the results return negative.

15.6.9 After quarantine, the animals shall be moved to their respective holding rooms, as requested by the PI/Researcher on the form.

15.7 DOCUMENTATION


15.8 APPENDICES

15.8.1 Appendix I: Animals Import Form

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Appendix I: Animals Import Form

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Title: SOP for Exporting Animals

Audience: Laboratory Animal Facility Manager, Attending veterinarian, Researchers / PIs


16.1.1 This document describes the procedure for exporting rodents to other local facilities or countries from the NTU’s Animal Research Facility.

16.1.2 The purpose is to ensure that the required documents and permits for exporting animals are properly obtained. It also ensures that the animals to be transported have proper care during the export.


16.2.1 Animals Export Form (ARF_ANIMALEXPORT01)

16.2.2 Animal Packing List

16.2.3 AVA Export Permit (for overseas export)

16.2.4 AVA Veterinary Certificate (for overseas export)


16.3.1 The abovementioned AVA documents can be found on the Agri-Food & Veterinary Authority of Singapore Website: www.ava.gov.sg.

16.3.2 The related AVA documents must be paid in full by the researcher / PI prior to the commencement of the export.

16.3.3 The PIs, facility managers and responsible veterinarians from both facilities must be informed of this export.

16.3.4 The courier that is selected by the researcher / PI to handle this export should advise the documents that are required on both sites for this export.

16.3.5 The animals to be exported must be checked and verified healthy by the attending veterinarian within 2 weeks prior to the export.

16.3.6 All logistics for this export has to be prepared by the researcher / PI.

16.4 DEFINITIONS

16.4.1 N.A.

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16.6 PROCEDURE

16.6.1 The researcher / PI shall source and select a courier service to handle this export. Subsequently, this request shall be made by filling and submitting the Animals Export Form to the NTU’s facility manager ensuring that the terms and conditions listed on the form are adhered to. This is subjected to approval by the facility manager and attending veterinarian.

16.6.2 The attending veterinarian shall use this form to locate and check the mice to be exported to verify if they are fit enough to be shipped. These mice should be checked again within 2 weeks prior to the export.

16.6.3 The researcher / PI shall apply and pay in full for the AVA Export Permit and the AVA Veterinary Certificate prior to the export. Once successfully applied, these shall be submitted to the facility manager for records.

16.6.4 The facility manager shall submit a copy of the health monitoring report to the consigning facility, upon request.

16.6.5 When the facility manager and the attending veterinarian approve the export, the Animals Export form shall be signed and returned to the researcher / PI and the consigning facility.

16.6.6 Upon approval and confirmation of the export date and time, the researcher / PI shall arrange and prepare all the logistics required for the export, including transporting crates, dividers, nectar gels and the animal packing list. This list is for the consigning facility to identify the animals that they will receive. Nectar gels may not be required for local exports.

16.6.7 Clean bedding and diet can be requested from the animal caretakers, ideally 1-2 days before the export.

16.6.8 The rodents shall be packed by the researcher/PI, together with sufficient nectar gels, bedding, and diet. The rodents shall be checked for any abnormalities once more. Wet diet may be provided for local exports.

16.6.9 The crates shall be properly labelled and sealed. They shall be passed to the courier via the usual exit route.

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16.6.10 In the event that further animal testing is required by the consignee’s

facility/country/governing authorities, or if recommended by the attending veterinarian, the cost of this test(s) shall be borne by the researcher/PI.

16.7 DOCUMENTATION


16.8 APPENDICES

16.8.1 Appendix I: Animals Export Form

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Appendix I: Animals Export Form

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Title: SOP for Animal Carcass Disposal

Audience: All animal handlers


17.1.1 This document describes the procedure for the safe disposal of animal carcasses at the NTU Animal Research Facility (ARF), in a manner that will not contaminate other animals, employees or the community.

17.1.2 The purpose is to ensure that the carcass disposal is compliant to the NACLAR guidelines and all regulations.


17.2.1 N.A.


17.3.1 Only mice and rats are housed at the NTU ARF.

17.3.2 The rodents housed at the ARF shall only be euthanized according to the methods described in the respective Animal Use Protocol forms.

17.3.3 The rodents shall only be euthanized in procedure rooms, never in animal holding rooms.

17.3.4 Researchers / Principal Investigators are responsible for recording the euthanized animals.

17.4 DEFINITIONS




17.4.4 PI – Principal Investigator


17.4.6 NACLAR Guidelines - National Advisory Committee on Laboratory Animal Research Guidelines


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17.6 PROCEDURE

17.6.1 Euthanised animals at Level 1

17.6.1.1 The carcass shall be placed in a biohazard bag which is available in all procedure rooms.

17.6.1.2 This bag shall be tied up or taped and transported to the refrigerator located in the clean area of the ARF Level 1. This carcass must be transferred to the chest freezer at the dirty area of level 1 within 48 hours.

17.6.1.3 Alternatively, the carcass can be transported to the chest freezer at the dirty area of level 1 and stored until it is due for collection by the designated carcass collector.

17.6.2 Euthanised animals at Level 2

17.6.2.1 Carcass shall be bagged in the same manner as 6.1.1. The bag shall be transported to the carcass freezer at the cage wash area.

17.6.2.2 The bags of carcass in the freezer shall be transported using a trolley to the chest freezer at the dirty area of level 1 at the end of every week.

17.6.3 Animals found dead

17.6.3.1 If an animal is found dead in a cage, the animal caretaker shall identify it if possible and record it on the cage card.

17.6.3.2 This shall also be recorded in the daily census sheet.

17.6.3.3 The researcher / PI shall be informed. They will then make a decision to discard the carcass or process it.

17.6.3.4 The carcass disposal is similar to 6.1 and 6.2, depending on the level that it was found.

17.7 DOCUMENTATION


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Title: SOP for Animal Research Facility Waste Management

Audience: All ARF Personnel


18.7.1 This document describes the procedure for safe management of the waste generated by the Animal Research Facility (ARF).

18.7.2 The purpose of this document is to ensure that all personnel, including the animal caretakers, PIs, researchers and officers are familiar with the disposal method and location of the different type of wastes produced at the ARF.


18.2.1 Revised Procedures for Waste Disposal

18.2.2 NTU Safety Manual for Biological Work


18.3.1 The personnel shall put on full PPE as described in the Procedure for Entry and Exit of ARF when handling wastes.

18.3.2 There are 2 units of Techniplast DS36 Bedding Disposal Station present in the NTU’s Animal Research Facility, one in Level 1 Dirty Area and another in Level 2 Cage Wash Area. These 2 units are serviced annually by ITS Science & Medical Pte Ltd and dependent on contract. They provide high protection against allergens and contaminants for the operator / animal caretaker.

18.3.3 Four main types of waste are generated by the ARF: General Waste, Biohazard Waste, Sharps and Animal Carcasses. Liquid Biohazard Waste is managed by researchers.

18.3.4 A summary of the waste management by ARF is shown in the table below:

Type of Waste Receptacle Double Bag? First Step Final Step

General Waste Trash Bag Not required Discard as normal waste

Normal Bedding Trash Bag Not required Discard as normal waste

Solid Biohazard Waste Biohazard Bag Yes Discard in Biohazard Bin

Collection by waste collector

Biohazard Bedding Biohazard Bag Yes Discard in Biohazard Bin


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Animal Carcasses Biohazard Bag Yes Store in Freezer Collection by

waste collector

Cytotoxic Waste Cytotoxic Bag Yes Discard in Cytotoxic Bin


Glass and Sharps Waste Sharps Bin Secure tightly Transport to SBS B1 for

disposal


Liquid Biohazard Waste (responsibility of

researcher)

Carboy Container with

GHS Label

Secondary containment

Transport to SBS B1 for

disposal N.A.

18.4 DEFINITIONS

18.4.1 Solid biohazard waste: Any form of non-sharp solids that have been contaminated with animal or human materials such as blood, tissue debris and any microbiological materials. They are autoclaved by the waste collector and disposed as normal waste.

18.4.2 Liquid biohazard waste: Bulk quantities of body fluids, media, microbiological cultures etc. These are autoclaved and disposed as normal waste.

18.4.3 Cytotoxic waste: biological waste contaminated with carcinogenic chemicals. These are incinerated by the waste collector before disposal.

18.4.4 Sharps waste: Any form of waste that includes an object that can puncture or lacerate the skin. It must be treated as biohazardous.


18.5.1 All ARF personnel

18.6 PROCEDURE

18.6.1 Non-Biohazard Bedding Disposal in Level 1

18.6.1.1 The technician shall push trolley with dirty cages to the Level 1 dirty area for bedding disposal from the dirty corridor.

18.6.1.2 The technician shall switch on the main power supply to the machine.

18.6.1.3 The technician shall press the green button on the control panel to activate the ventilation.

18.6.1.4 When cleaning dirty bedding for BSL1 cages, the technician shall place normal trash bag below the machine. When cleaning dirty bedding for BSL2 cages, the technician shall place a biohazard bag below the machine. Cleaning of these cages shall be done on separate occasions to avoid mixing the bedding.

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18.6.1.5 The blower shall be left to run for at least 15 minutes before usage. This is

to ensure that it has achieved the programed air flow.

18.6.1.6 The personnel shall proceed to discard dirty bedding from a cage in the machine by scrapping the dirty bedding with a scraper, allowing the dirty bedding to collect below. Movements in and out of the machine shall be swift and smooth to minimize disruption to the air barrier.

18.6.1.7 The normal trash bag shall be removed from the bin when it is about 70% full. The personnel should either tie a knot to the bag or tape the bag to prevent spillage. If the bag is torn, it shall be placed into a new bag and secured in the same manner.

18.6.1.8 Normal waste shall be transported to the carcass disposal area. The designated cleaner will open the shutter from the outside to transfer the bags of normal trash to the large garbage bin situated just outside. The designated waste collector will collect the trash daily.

18.6.2 Biohazard Bedding Disposal in Level 2

18.6.2.1 The technician shall arrange the dirty cages in the Cage Wash Area to prepare them for bedding disposal.

18.6.2.2 The technician shall follow the steps described from 6.1.1 to 6.1.7.

18.6.2.3 For biohazard waste bags, the bag shall be taped with autoclave tape, double-bagged and taped in the same manner.

18.6.2.4 The bags of biohazard waste shall be transported down to Level 1 using a trolley via the elevator then disposed into the biohazard bin at the unloading area.

18.6.2.5 The technician shall contact Sembcorp for waste collection when the bin is 50% full.

18.6.3 General Waste Disposal

18.6.3.1 General wastes are collected in normal black trash bags. General wastes include empty glove boxes, torn but uncontaminated gloves, external packaging etc.

18.6.3.2 In Level 1, the technician shall collect the general waste from each bin and transport them to the dirty area via the dirty corridor.

18.6.3.3 The technician shall then proceed with step 6.1.8 to discard the trash.

18.6.3.4 In Level 2, the technician shall transport the general waste to Level 1, and follow the steps described in 6.2.5 and 6.2.6.

18.6.4 Biohazard Waste and Cytotoxic Waste Disposal

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18.6.4.1 All personnel shall discard biohazard and cytotoxic waste into the

respectively labelled bags (biohazard waste bags are yellow with a biohazard symbol while cytotoxic waste bags are purple with the cytotoxic symbol).

18.6.4.2 When the bags are about 70% full, the personnel shall seal it with autoclave tape.

18.6.4.3 The personnel shall then place it in another biohazard or cytotoxic bag respectively and seal it with autoclave tape.

18.6.4.4 Biohazard and cytotoxic waste from Level 1 and 2 shall be brought to the unloading area and dispose them into the respective bins.

18.6.4.5 When the bins are 50% full, the technician shall contact the waste collector for collection.

18.6.5 Animal Carcass Disposal

18.6.5.1 Animal carcasses shall be disposed into biohazard bags, sealed with autoclave tape then double-bagged and sealed in the same manner.

18.6.5.2 These bags shall be transported to the carcass disposal area via the dirty corridor.

18.6.5.3 The carcasses are stored in the freezer. When they freezers are 70% full, the technician shall contact the waste collector for collection.

18.6.6 Glass and Sharps Waste Disposal

18.6.6.2 All personnel shall refer to NTU OHS’s Guideline on Handling of Sharps for more information.

18.6.6.3 All glass and sharps must be disposed into the sharps bin located in the procedure rooms.

18.6.6.4 All users shall avoid discarding non-sharps, such as the syringes, syringe needle caps, syringe packaging etc. into the sharps bin.

18.6.6.5 The sharps waste shall not exceed the line indicated on the sharps bin, i.e. about three-quarters full.

18.6.6.6 The sharps bin shall be capped shut when it is sufficiently full.

18.6.6.7 The technician shall then place the bin in a biohazard bag and seal it with autoclave tape.

18.6.6.8 The technician shall transport this to the BSL2 laboratory to autoclave.

18.6.6.9 After autoclaving the sharps bin, the technician shall transport the bins to SBS B1 for disposal.

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18.6.7 Liquid Biohazard Waste and Liquid Chemical Waste Disposal

18.6.7.1 Researchers are responsible for their own liquid biohazard waste.

18.6.7.2 All carboys must be labelled with a proper GHS label, including the type of waste and PI’s name.

18.6.7.3 All carboys must be placed in secondary containment.

18.6.7.4 The carboys shall be filled to about 80%, and subsequently transported to SBS B1 for disposal.

18.7 DOCUMENTATION

18.7.1 All wastes disposal, except general waste, must be recorded in the Waste Document (Appendix I).

18.8 APPENDIX

18.8.1 Appendix I: Waste Document Appendix I

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Ref. No.: NTU-ARF-SOP-19 Version: 1

Date of Issue: 18-Feb-15 Next Review Date: 18-Feb-18

Title: Emergency Response Plan of Animal Research Facility

Audience: All staff members, users and visitors.


19.1.1 This document serves as a basic plan in case of emergencies or disasters that occur at the Nanyang Technological University (NTU) Animal Research Facility (ARF). This is a supplementary document to the Emergency Response Plan (ERP) for School of Biological Sciences (SBS) NTU.

19.1.2 The list of fire wardens and OFA of the ARF can be found at the main entrances of the building and on the first-aid kits respectively.

19.1.3 The first priority in any emergency situation is the prevention of injury and the immediate attention and assistance to the injured person(s) followed by research animal(s).


19.2.1 Emergency Response Plan for School of Biological Sciences, NTU.


19.3.1 The premise described in this document is located at

i. Animal Research Facility ii. 60 Nanyang Drive iii. Singapore 637551 iv. Laboratory Animal Facility Manager: Ms Lee Shok Li v. Tel: 6316 2921

19.3.2 SBS ARF Emergency contact information:

SBS Emergency Response Team………………………………….63162803, 65148392

Major Emergency……..……………………………………………...6790 5200 (24hr)

NTU Medical Centre………………………………………………….67936828/ 67936974

Police……………………………………………………………...……999

Fire and Ambulance……………………………………………..……995

List of full-time employees at Animal Research Facility:

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No. Staff Name Email Number Location Position

1 Lee Shok Li [email protected]

63162921 97860585

SBS-01-AH11 Assistant Manager,

Animal Research Facility

2 Panchatcharam Manisekaran

[email protected] 63162944/5 ARF Level 1 Executive, Animal Research Facility

3 Kalyanasundaram

Nageswari [email protected] 63162944/5 ARF Level 1 Executive,


4 Manickam Vennila [email protected] 63162944/5 ARF Level 1 Assistant Executive, Animal Research Facility

5 Ravichandran

Annamalai [email protected] 63162944/5 ARF Level 2

Assistant Executive, Animal Research Facility

6 Husazli Bin Hussin [email protected] 63162944/5 ARF Level 2 Assistant Executive, Animal Research Facility

7 Myo Min (Lionel) [email protected] 63162810 SBS-01n-21 Project Manager, Safety

(at SBS GO)

19.3.3 The different types of emergencies that can trigger this procedure are summarized below:

No. Emergency Condition Possible Scenario *Control Measure

(i) Fire Emergency

• Flammables, open flame & Ignition

• Chemical reaction • Electricity overload • Fire alarm

• SBS ERP • Proper use / storage of flammable

materials • Fire Evacuation protocol

(ii) Biological Spill • Spill / Clean up

Spill in BSC, centrifuge, etc. • SBS ERP • Decontaminate / Clean up

(Refer to: www.ntu.edu.sg/ohs)

(iii) Chemical/Radioactive Spill

• Spills during experiment • SBS ERP • OHS Spill Guideline

(Refer to: www.ntu.edu.sg/ohs)

(iv) Explosion • Centrifuge/ Autoclave (Failure

to observe SOPs) • Chemical reaction

• SBS ERP • Equipment operation manual

(v) Medical Emergency

• Splash to eyes • Cuts, Abrasion, skin contacts • Thermal / chemical burn • Exposure to chemical with

hazard

• SBS ERP • Use of PPE • Medical Emergency protocol • First aid facility

(vi) Power Outage • Overloading of power socket • SBS ERP

(vii) Personnel Injury

• Animal bites, thermal burn from autoclave, ergonomics injury

• Responsible Care and Use of Laboratory Animals (RCULA) Training

• Use of PPE • Medical Emergency protocol • First aid facility

(viii) Protests against Animal Research

• Animal Activist threats • Notification of the authority

(ix) Natural Disasters • Flooding caused by heavy downpour or backflow from pipes

• Relocation of animals

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19.3.4 There are 2 emergency exits located on each level of the Animal Research

Facility (ARF).

19.3.5 Exit routes must be free of flammable or combustible materials.

19.4 DEFINITIONS

19.4.1 N.A.




19.5.3 Researchers

19.5.4 Visitors


19.6 EMERGENCY RESPONSE PROCEDURE FOR ARF

19.6.1 Fire Emergency

19.6.1.1 Refer to the Emergency Response Plan for SBS and notify the people described in the actions for office hours and post-office hours.

19.6.1.2 Activate the fire alarm and notify the people in the vicinity about the emergency. Notify Laboratory Animal Facility Manager (LAFM) and the Fault Reporting Centre (FRC) immediately.

19.6.1.3 Evacuate the building via the nearest exit immediately. The Fire Wardens shall notify and evacuate everyone by knocking and calling out on each door and corridor. All personnel shall proceed to the Assembly Point (The QUAD) and wait for instructions from the Liaison Officer.

19.6.1.4 First Aiders shall attend to the injured at the Assembly Point.

19.6.1.5 The Fire Safety Manager shall confirm if it is safe to return to the workplace after the emergency has been mitigated or investigated.

19.6.2 Biological, Chemical/Radioactive Spill

19.6.2.1 Refer to the ERP for SBS for spill management.

19.6.2.2 Spill kits are available on both levels of the ARF.

19.6.2.3 All animal caretakers are trained in spill management.

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19.6.2.4 Discard the bags containing the spill accordingly. Biohazard spills shall be

autoclaved in the BSL2 laboratory before disposal. Chemical spills shall be transported to the Basement 1 of SBS for disposal.

19.6.3 Bomb Threats / Explosions

19.6.3.1 Refer to the ERP for SBS for appropriate actions to take.

19.6.4 Medical Emergency and Animal Bites

19.6.4.1 Refer to the ERP for SBS for appropriate actions to take for medical Emergencies

19.6.4.2 All personnel shall be wearing appropriate personal protective equipment, PPE, at all times while working in the ARF.

19.6.4.3 First Aid kits are present on both levels of the facility. Stretcher is also available. There are 2 designated Occupational First Aiders in the building.

19.6.4.4 Please refer to the SOP for Animal Bites, Scratches and Animal-related Injuries.

19.6.5 Power Outage

19.6.5.1 Refer to the ERP for SBS for appropriate actions to take for power outage.

19.6.5.2 The emergency lights will be lit, and the exit signs should be visible along all corridors. The emergency generator will supply power to the air handlers and fans.

19.6.5.3 Personnel shall remain calm and proceed to a lit area or exit the room carefully.

19.6.5.4 Personnel shall follow the exit lights to exit the facility.

19.6.5.5 Call and report the outage to the Campus Security, as described by the ERP for SBS.

19.6.5.6 As soon as the power returns, the animal caretakers shall proceed to all the animals and check that the individually ventilated cage (IVC) systems are working normally. Any animals in distress shall be monitored closely. If treatment is required, contact the Attending Veterinarian.

19.6.5.7 All abnormalities shall be reported to the LAFM.

19.6.6 Protests against Animal Research

19.6.6.1 In the event of a protest or animal activist threat, the Campus Security shall be notified immediately. They will decide what actions to take in response of the threat.

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19.6.7 Natural Disasters

19.6.7.1 If a flood from natural disaster or burst pipe occurs, notify the FRC, Campus Security and the LAFM immediately.

19.6.7.2 The extent of the flood shall be checked immediately. If the flood is ankle deep in the animal holding rooms and is worsening, the racks shall be unplugged from the systems and moved out of the facility via the emergency exits.

19.6.7.3 The LAFM shall inform the A*Star Biological Resource Centre to request for temporary holding rooms to hold the animals. The animals shall be checked as soon as they have been relocated.

19.6.7.4 The animals shall be held at BRC while the facility is thoroughly cleaned and sanitized.

19.6.7.5 If the natural disaster affects the ARF food and bedding supplies, they shall be inspected for damage and usability. The local supplier shall be contacted for immediate delivery of supplies. Spoilt food shall be discarded.

19.7 DOCUMENTATION


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Title: SOP for Animal Bites, Scratches and other Animal-Related Injuries

Audience: All animal handlers.


20.7.1 This document describes the management of animal bites, scratches and animal-related injuries. It is intended for all personnel who have to handle animals, or who may be exposed to animal bodily materials.

20.7.2 All Principal Investigators are responsible for ensuring that their staff members are aware of the risks involved with working with animals. This SOP applies to all users of the NTU Animal Research Facility (ARF).

20.7.3 Only rodent-related injuries are covered in this protocol.


20.2.4 Emergency Response Plan for School of Biological Sciences, NTU.


20.3.1 An animal bite is defined as having ones skin punctured by the animal’s teeth and having the animal saliva coming in contact with the exposed skin. The animal’s claws may also puncture one’s skin and come into contact with human tissue, eyes or mucous membranes. Bites and scratches are considered as physical injuries, and carry the potential of contracting zoonotic diseases and/or allergic reactions.

20.3.2 Only mice and rats are housed at the NTU ARF.

20.3.3 All personnel and animal caretakers must be vaccinated with tetanus and Hepatitis B.

20.3.4 There are 3 first-aid kits located in the building:

a. Laboratory Animal Facility Manager’s Office b. Level 1 General Store c. Level 2 Dirty Corridor

20.3.5 In case of medical emergencies caused by animal-related injuries, the list of SBS ARF Emergency contact information can be found below:

SBS Emergency Response Team……………………..…..…6513 7652

Major Emergency: ODFM …………………………………..6790 5200 (24hr)

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NTU Medical Centre…………………………………………….67936828

Fire and Ambulance…………………………………………….995

List of full-time employees at Animal Research Facility:

No. Staff Name Email Number Location Position

1 Lee Shok Li [email protected]

63162921 97860585 SBS-01-AH11

Assistant Manager, Animal Research Facility

2 Panchatcharam

Manisekaran [email protected] 63162944/5 ARF Level 1

Executive, Animal Research Facility

3 Kalyanasundaram

Nageswari [email protected] 63162944/5 ARF Level 1 Executive,


4 Manickam Vennila [email protected] 63162944/5 ARF Level 1 Assistant Executive,


5 Ravichandran

Annamalai [email protected] 63162944/5 ARF Level 2 Assistant Executive,


6 Husazli Bin Hussin [email protected] 63162944/5 ARF Level 2 Assistant Executive, Animal Research Facility

7 Myo Min (Lionel) [email protected] 63162810 SBS-01n-21 Project Officer, Safety

(at SBS GO) Table 1* Occupational First-Aiders

20.3.6 PPE is provided in the facility. New gloves are provided in all animal holding rooms and procedure rooms. Animal handlers shall put on extra gloves whenever deemed necessary.

20.3.7 All animal handlers must have attended the Responsible Care and Use of Laboratory Animals (RCULA) course and must be working on an IACUC-approved protocol before they are allowed to work at the Animal Research Facility.

20.4 DEFINITIONS


20.4.2 ODFM – Office of Development and Facilities Management


20.4.4 RCULA – Responsible Care and Use of Laboratory Animals course

20.4.5 OFA – Occupational First-Aider




20.5.3 Researchers

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20.5.4 Visitors


20.6 PROCEDURE

20.6.1 Immediately flush a bite or scratch wound with plenty of soap and water.

20.6.2 Locate the First-Aid kit. If the injured personnel are trained in first-aid and able to apply it onto themselves, he or she shall proceed to do so. Otherwise, contact the OFA for assistance if the incident happened during office hours.

20.6.3 If the incident happened after office hours, apply first-aid if the injured personnel are trained and if it is a small wound. If further medical assistance is required, the injured personnel shall call the NTU Medical Centre or ambulance.

20.6.4 The injured personnel shall report the incident to the LAFM as soon as possible. The LAFM will complete an incident report which will be reviewed and determine if further actions shall be taken. The follow-up actions may include review of SOP, retraining or review of risk assessments.

20.7 DOCUMENTATION


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Title: SOP for Preparation of Acidified Drinking Water for Laboratory Rodents



21.8.1 This document describes the procedure for the safe preparation of acidified drinking water for the laboratory rodents at SPF Level 1.

21.8.2 The purpose of this is to ensure that the animal caretakers are aware of the hazards and risks involved in handling hydrochloric acid, and the safety measures to take.


21.2.1 Safety Data Sheet for Hydrochloric Acid, 37%, Fuming


21.3.1 The purpose of acidifying the laboratory animal drinking water continuously is for disinfection. Acidified water has a bactericidal effect, against Pseduomonas aeruginosa and a few other Gram-negative bacteria. This prevents the spread of opportunistic pathogens between animals.

21.3.2 Merck’s Hydrochloric acid fuming 37% is added into the drinking water for the laboratory rodents.

21.3.3 The pH of the acidified drinking water is approximately pH 2.5 – pH 3.

21.3.4 Some microorganisms may still grow in the water but may not multiply.

21.3.5 The animal caretakers must be familiar with the Safety Data Sheet of the HCl.

21.3.6 Full PPE, safety goggles and covered shoes must be worn at all times while working with HCl.

21.3.7 The dispensing of HCl must be done in the fume hood with absorbent material in the vicinity. The spill kit must be in the same room.

21.3.8 Concentrated HCl must only be handled in the fume hood.

21.3.9 HCl is stored in the acid cabinet, located below the fume hood at Procedure Room 2.

21.3.10 Always add acid into water, and not water into acid!

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21.4 DEFINITIONS






21.6 MATERIALS

21.6.1 Hydrochloric Acid, 37%, Fuming

21.6.2 Automatic pipette pump

21.6.3 Serological Glass Pipette, 25 mL

21.6.4 1 L of water in Nalgene glass bottle

21.6.5 Water tub

21.6.6 Cleaned water bottles with sipper tubes

21.7 PROCEDURE

21.7.1 Switch on the fume hood and ensure that it is working.

21.7.2 Place a piece of absorbent tissue paper on the working surface of the fume hood.

21.7.3 Place the bottle of water, glass pipette and pump in the hood.

21.7.4 Using both hands, retrieve the bottle of HCl from the acid cabinet located below the fume hood and place it on the working surface.

21.7.5 Using the pump and pipette, aliquot the acid slowly and carefully dispense it into the bottle of water in similar motion. The amount of acid to aliquot are as follows:

21.7.6 For a final volume of 70L, aliquot 21.0 mL of acid into the 1L bottle of water. A pH of about 2.5 – 3.0 is achieved.

21.7.7 Screw the cap securely onto the glass bottle and bring it to the water preparation area.

HCl Water 1.5 mL 5 L 9.0 mL 30 L

21.0 mL 70 L

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21.7.8 With the water tub placed on a trolley, fill the water tub with tap water to the

desired level and pour the acidified bottle of water into the tub slowly and in a circular motion.

21.7.9 Arrange clean, empty water bottles on a trolley.

21.7.10 Dunk the water bottles into the water tub and fill them with the acidified water, carefully to avoid splashing.

21.7.11 Dry them on a towel and place them on another trolley.

21.7.12 Cap the filled water bottles with the sipper tubes and place the bottles in the box for autoclaving later.

21.7.13 Repeat until all the boxes are filled with water bottles.

21.7.14 Cover the boxes and transport them to the autoclave area using a trolley.

21.7.15 Push them into the autoclave and start the autoclave cycle.

21.7.16 After the autoclave cycle, remove the box and allow it to cool. These water bottles are now ready for use.

21.8 DOCUMENTATION


21.9 REFERENCES

21.9.1 Edstrom Industries, 2003. Drinking Water Acidification [online]. Wisconsin: Edstrom Industries, Inc. Available: http://www.edstrom.com/file.aspx?DocumentId=69 [Accessed 18 February 2015].

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