HB, Gel Staining

um

Processor Plusautomated Western blot processing and gel staining

user manual

DNA/Protein Labeling, Hybridization & Detection

80-6446-89 Rev B/5-99

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1 Introduction to Processor Plus . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

Optional printing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1Protocol key . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

2 Instrument Components . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2

Connecting the Processor Plus to a serial printer . . . . . . . . . 4

3 Navigating the Operating Software . . . . . . . . . . . . . . . . . . . . . . . . 5

Reviewing protocols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6Reviewing the Setup parameters . . . . . . . . . . . . . . . . . . . . . . . . 7

4 Setting Up for Blot Processing . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

Configuring the blot processing tray and pump tubing . . . . . . 8

Vent line . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

Leveling the tray . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

Calibrating the pump . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

5 Operating Instructions for Blot Processing . . . . . . . . . . . . . . . . . 13

Prepare for blot processing . . . . . . . . . . . . . . . . . . . . . . . . 13

Starting at the beginning of the protocol . . . . . . . . . . . . . . . . . . 14Starting at a particular step in the protocol . . . . . . . . . . . . . . . . 15Interrupting a protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

Cleaning protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

6 Editing Blot Processing Protocols . . . . . . . . . . . . . . . . . . . . . . . . 17

Editing a protocol name . . . . . . . . . . . . . . . . . . . . . . . . . . 17

Editing step variables . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17

Port In . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18Minutes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18Multiplier . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18Hold, Beep and Rock (HBR) . . . . . . . . . . . . . . . . . . . . . . . . . . 18Editing when the Processor Plus is not running a protocol . . . . . 19Adding or inserting a step . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19Deleting a step . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20Editing a protocol in progress . . . . . . . . . . . . . . . . . . . . . . . . . 20

7 Setup Parameters for Blot Processing . . . . . . . . . . . . . . . . . . . . . 22

Setup 1: Manual pump operation . . . . . . . . . . . . . . . . . . . . . . . 22Setup 2: Tray size . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23Setup 3: Tray level . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23Setup 4: Display protocol volume . . . . . . . . . . . . . . . . . . . . . . . 23Setup 5: Print protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24Setup 6: Print volume of protocol . . . . . . . . . . . . . . . . . . . . . . . 24Setup 7: Pump calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . 25

8 Setting Up for Gel Staining . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26

Configuring the gel staining tray and pump tubing assembly 27

Leveling the tray . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27

Calibrating the pump . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28

Hoefer Processor Plus

9 Operating Instructions for Gel Staining . . . . . . . . . . . . . . . . . . . . 30

Prepare for gel staining . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30Starting at the beginning of the protocol . . . . . . . . . . . . . . . . . . 30Starting at a particular step in the protocol . . . . . . . . . . . . . . . . 31Interrupting a protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31

Cleaning protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 32

10 Editing Gel Staining Protocols . . . . . . . . . . . . . . . . . . . . . . . . . . 33

Editing a protocol name . . . . . . . . . . . . . . . . . . . . . . . . . . 33

Editing step variables . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33

Port In . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34Minutes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34Port Out . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34Hold, Beep and Rock . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 34Editing when the Processor Plus is not running a protocol . . . . . 35Adding or inserting a step . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36Deleting a step . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36Editing a protocol in progress . . . . . . . . . . . . . . . . . . . . . . . . . 36

11 Gel Staining Setup Parameters . . . . . . . . . . . . . . . . . . . . . . . . . . 38

Setup 1: Manual pump operation . . . . . . . . . . . . . . . . . . . . . . . 38Setup 2: Tray size . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39Setup 3: Volume delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39Setup 4: Tray level . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39Setup 5: Print protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 40Setup 6: Pump calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . 40

12 Using the protocol key . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

13 Troubleshooting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 43

14 Care and Maintenance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

Running the cleaning protocol . . . . . . . . . . . . . . . . . . . . . 47

Staining trays . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

Chemical Resistance . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48

Cleaning the splash guard . . . . . . . . . . . . . . . . . . . . . . . . 48

Replacing the tray gasket on the staining tray . . . . . . . . . . . 48

Pump Care and Maintenance . . . . . . . . . . . . . . . . . . . . . . 49

A Pre-programmed Blot Processing Protocols . . . . . . . . . . . . . . . . 54

B Pre-programmed Staining Protocols . . . . . . . . . . . . . . . . . . . . . . 58

C Blot Process Protocol Worksheet . . . . . . . . . . . . . . . . . . . . . . . . 63

D Gel Stain Protocol Worksheet . . . . . . . . . . . . . . . . . . . . . . . . . . 64

E Specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65

Customer Service Information . . . . . . . . . . . . . . . . . . . . . . . . . . 66

Ordering Information . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67

Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69


S a f e t y W a r n i n g s a n d P r e c a u t i o n s

Safety Warnings and Precautions

WARNING For Research Use Only. Not recommended or intended for diagnosis of disease in humans or animals. Do not use internally or externally in humans or animals.

WARNING Not recommended or intended for use with radioisotopes.

We recommend that this product is handled only by those persons who have been trained in laboratory techniques and that it is used in accordance with the principles of good laboratory practices, as all chemical should be considered as potentially hazardous. When handling chemical reagents, wear suitable protective clothing such as laboratory overalls, safety glasses and gloves. Avoid chemical contact with skin or eyes. In case of contact with skin or eyes, wash the affected area immediately with water.

If this equipment is used in a manner not specified by the manufacturer, the protection provided may be impaired.

Plug the instrument into a properly grounded outlet. Always disconnect the power cord before servicing.

Avoid spilling liquids on the body of the instrument.

Ensure that the vents at the side and bottom of the instrument are not blocked.

Use only the supplied reagent line assemblies. Detaching the rigid opaque tubing or using different reagent lines invalidates the pump calibration.

Periodically check that no liquid is accumulating in the tray support.

Only accessories and parts approved or supplied by Amersham Biosciencesmay be used for operating, maintaining, and servicing this product.


I n t r o d u c t i o n t o P r o c e s s o r P l u s

1 Introduction to Processor Plus

The Hoefer Processor Plus automates fluid delivery and timing steps for both membrane processing and polyacrylamide gel staining.

The Processor Plus base unit consists of:

• the electronic and mechanical parts, encased in a metal chassis. The instrument control panel—an LCD and keypad—is on the front of the unit.

• A peristaltic pump which works in conjunction with the 10-port valve to transfer fluid from reagent bottles to the tray. Both the pump and the 10-port valve are on the back of the unit.

• Memory on the Processor Plus that can hold up to 24 protocols: 10 for membranes and 14 for gels. Each protocol can hold up to 30 steps.

A tray and lid sit on top the base unit to hold either the gels or membranes during staining or blot processing.

The blot processing tray option includes a tray support, two Mini trays, one Standard tray and one blot processing lid that fits either tray.

The gel staining tray option includes a tray support and a choice of two tray sizes—125 ml and 250 ml—each with its own lid and tray support.

Optional printing

You can connect the Processor Plus to a serial printer via a 9-pin cable and adaptor (code no. 80-6427-70). See page 4 for instructions. When a serial printer is connected to the Processor Plus while a protocol is running, a paper report of each step in a protocol is generated, including:

• the volume of reagents used in each step

• the time, in minutes, for each step

• a validation report form, with spaces for sample ID, date, and operator

When the Processor Plus is not in use, you can use the Setup menus to print all the steps in a protocol or calculate the volume of each reagent needed.

Protocol key

The protocol key is a removable device that can store one protocol. You can use the protocol key to record a protocol, then remove the protocol key and store it in a safe place. Use the key to repeatedly run the stored protocol. The protocol on the protocol key does not change unless you overwrite it.

For more information on using the protocol key, see page 41.

Hoefer Processor Plus � p 1

Instrument Components

2 Instrument Components

Fig 2-1 Processor Plus components and accessories

6

7

8

95

3

1

10

11

4

2

12

No. Component Function

1 Rocker finger Provides the rocking motion for agitating fluid.

2 Pivot ball assemblies (2)

Fit into the sockets on the bottom of the tray support.

3 LCD Screen Displays protocol status and program steps.

4 Keypad Provides keys to program protocols and control the Processor Plus.

5 Protocol key slot Accepts the protocol key

6 Reagent lines Connects reagent bottles to 10-port valve.

7 10-port valve Selects reagent line to connect to peristaltic pump.

8 Peristaltic pump Pumps fluids from the 10-port valve into the tray and out of the tray to waste or bottle.

9 Pump tubing Carries reagents through peristaltic pump.

10 Protocol key Stores one protocol in removable memory.

11 Spirit level Assists in leveling trays.

p 2 � Hoefer Processor Plus

I n s t r u m e n t C o m p o n e n t s

Tray support. Fits onto the pivot ball assemblies on top of the metal chassis to

hold the tray and its lid.

Blot processing trays (two sizes). The Mini tray has four chambers, each holds

a maximum of 50 ml for a 9 × 9.5 cm membrane. The Standard tray has two chambers, each holds a maximum of 125 ml for a 16 × 16 cm membrane. Both trays are disposable.

Blot processing tray lid. A manifold on top of the blot processing lid delivers

reagents to the surface of the membranes. Valves on the manifold control delivery to chambers in the trays.

Staining trays (two sizes). Made of PTFE-coated stainless steel, the trays are

inert to most staining solutions. Two sizes are available: 19 × 29 and 29 × 35 cm.

Glass lid for gel staining (two sizes). Lids prevent spills and minimize exposure

to staining reagents while allowing observation of the staining process. Two sizes are available: 19 × 29 and 29 × 35 cm.

Coated magnets (4). When placed on gel staining tray, magnets prevent gels

from sliding over one another and from coming in contact with the splash guard. Placed on the corners of gels with plastic backing, they help keep gels submerged.


Instrument Components

Connecting the Processor Plus to a serial printer

An RS232C serial port with a 9-pin male D-sub connector is located on the bottom of the Processor Plus (Fig 2-2). A 9-pin cable and an adaptor for the serial printer connection is available in a serial printer cable kit (code no. 80-6427-70).

The signal and pin number assignments on the Processor Plus serial port are:

The serial printer must have the following settings:

To connect the Processor Plus to the serial printer:

1. Attach the 25-pin male adaptor to the port on the serial printer.

2. Connect one end of the 9-pin cable to the RS232C serial port on the

Processor Plus and the other end to the adaptor on the serial printer. See

Fig 2-2.

Fig 2-2 Locating the RS232C serial port

An example of a protocol printout

Pin 2 Transmit

Pin 3 Receive

Pin 5 Ground

Other pins not connected

Baud rate 1200

Data bits 8

Stop bit 1

Start bit 1

Parity None

Flow control None

RS232C serial port

9-pin cable connector


N a v i g a t i n g t h e O p e r a t i n g S o f t w a r e

3 Navigating the Operating Software

The LCD on the Processor Plus displays the current status of the instrument.

When you turn on the power switch, located on the left side of the instrument, a self-diagnostic cycle runs for approximately one minute. This cycle tests all circuits and moving parts. During the self-diagnostic cycle, the pump starts and stops, the instrument emits a “beep,” and the tray rocks.

If any component test fails, the self-diagnostic cycle stops and a message on the screen indicates the source of the fault. Press START to skip to the next test. Report component test failures to your nearest Amersham Biosciencesservice representative.

Once the diagnostic cycle is successfully completed, the screen displays the name of the last protocol run on the Processor Plus. Press STOP to return to the Main menu.

The keypad (Fig 3-1) contains three types of keys: Program, Arrow and Protocol.

Table 3-1 Keypad Keys

Fig 3-1 The Keypad

The Main menu is the entry to all protocols and menus on the Processor Plus. Use the keypad to navigate from the Main menu to either blot processing or gel staining protocols (see Fig 3-3).

Fig 3-2 The Main menu on the LCD

Type Key Names Function

Program EDIT/SET, ADD, DEL Edit protocols and the variables in each step of a protocol.

Arrow !, ", �, � Move cursor and change values on the LCD screen.

Protocol START, STOP, PAUSE Control protocols and setups.

Protocol keys

Arrow keys

Program keys

-> Process blot Stain gel


Navigating the Operating Software

Reviewing protocols

From the Main menu, choose to see either blot processing protocols or gel staining protocols.

Fig 3-3 Navigating through the protocols

1. From the Main Menu, press � or � to switch between the set of Process blot

and Stain gel protocols.

The arrow on the LCD screen indicates which protocol set is selected.

2. Press START to see the first protocol in the selected protocol set.

Press STOP to return to the Main menu.

3. Press � or � to review all the protocols in the protocol set.

4. Press EDIT/SET to see the first step in a selected protocol.

5. Press � or � to review all the steps in the protocol.

Press STOP to return to the list of protocols.

• For more on using the blot processing protocols, go to page 8.

• For more on using the gel staining protocols, go to page 26.

-> Process blot Stain gel � or �

Process blot-> Stain gel

Blot Protocol 1 ECL Plus

START

Gel Protocol 1 DNA Silver

START

1

2

Blot Protocol 2 ECL


Press � Press �3

STOP STOP

Press � Press �

(Step 1 in Gel Stain Protocol)

(Step 1 in Blot Process

EDIT/SET

STOP

(Step 2 in Blot Process

Press �

Press �

4

5

(Step 2 in Gel Stain Protocol)

EDIT/SET

STOP

Press �

Press �

Gel Staining ProtocolsBlot Processing Protocols

STOP STOP


N a v i g a t i n g t h e O p e r a t i n g S o f t w a r e

Reviewing the Setup parameters

The Processor Plus has two Setup menus, one for blot processing and one for gel staining (see Fig 3-4). Use the parameters in the Setup menus to:

• select a tray size

• level the tray

• set the volume of reagent pumped (for gel staining, only)

• calibrate the pump

• print a protocol

• print or display the total volumes of reagents used in the protocol (blot processing, only)

• manually operate the pump

Fig 3-4 Navigating to the Setup menus

1. From the Main Menu, press � or � to switch between the set of Process blot

and Stain gel protocols.

The arrow on the LCD screen indicates which protocol set is selected.

2. Press START to see the first protocol in the selected protocol set.

Once you have selected the appropriate protocol set, you can go to the Setup menu from any protocol.

3. Press and hold EDIT/SET until the Setup menu appears on the LCD.

Note You cannot use the Setup menus when the Processor Plus is running a protocol.

4. Press � or � to review the Setup parameters for the selected protocol set.

5. Press STOP to return to the protocols.

-> Process blot Stain gel � or �

Process blot-> Stain gel

1

Blot Protocol 1 ECL Plus

START


START

2STOP STOP

Gel Staining ProtocolsBlot Processing Protocols

3

STOP

Press and hold EDIT/SETPress and hold EDIT/SET

(Setup 1 in Blot processing)

(Setup 1 in Gel staining)

(Setup 2 in Blot processing)

(Setup 2 in Gel staining)

Press �

Press �

Press �

Press �

STOP

4

5

STOP STOP


Setting Up for Blot Processing

4 Setting Up for Blot Processing

The Hoefer Processor Plus allocates memory for ten blot processing protocols, five of these are pre-programmed (Table 4-1). See page 17 for directions on editing protocols.

The Processor Plus has two trays for blot processing (Table 4-2). One lid fits either tray.

Table 4-1 Pre-programmed blot processing protocols

Table 4-2 Tray Sizes for blot processing

Configuring the blot processing tray and pump tubing assembly

The lid assembly (Fig 4-1) on the membrane processing tray has:

• an inlet manifold in the centre

• a drain tube at the rear corner

• four valves for controlling reagent delivery to chambers

Fig 4-1 The membrane processing lid

The tubes from the lid assembly connect to the peristaltic pump. The tube from the inlet manifold connects to the inner peristaltic pump. The drain tube in the corner of the lid connects to the outer peristaltic pump. See Figure 4-2 on page 9.

Protocol Name Applications

1 ECL Plus ECL Plus Detection Kit

2 ECL ECL Detection Kit

3 Standard Immunodetection with 1° & 2° antibodies

4 Enhanced Immunodetection with 1°, 2° antibodies and avidin compounds

5 Clean Rinses reagents out of tubings, valves and ports

Tray Number of membranes

Size of membranes (cm)

Reagent Capacity(ml /chamber)

MiniStandard

1–4 1 to 2

9 × 9.516 × 16

10–50 25–125

Tray drain tube

Inlet manifold tube

Lid manifold


S e t t i n g U p f o r B l o t P r o c e s s i n g

The Quick-Fit couplings on the peristaltic pump tubing have different dimensions, keyed to the appropriate tubing connections on the trays. When the appropriate male and female couplings are pressed together, you can hear a click as they connect.

Fig 4-2 Configuration of membrane processing tubes and tray

Waste tube. Place the end of the waste tube into the top of a large waste bottle or

suspend it over the edge of the sink. A vented waste bottle cap (38-430), provided with blot processing trays, fits a narrow-mouth Nalgene® bottle. If you use a waste bottle that does not accept this cap, tape the end of the waste tube to the top of the waste container. Do not let the end of the waste tube become submerged in liquid.

Vent line

Important For membrane processing, reagent line 0 (zero) is reserved for air venting. Do not use reagent line 0 to deliver reagents.

The presence of detergent in some reagents creates bubbles. As a result, small amounts of reagent may get into the vent line, but this does not affect membrane processing quality. However, spray from these reagent drops escapes out the vent line when the Processor Plus is operating. Suspend the end of the vent line over a sink or large beaker to contain escaping spray.

After configuring the tubing and tray, level the tray and calibrate the pump.

Leveling the tray

For reliable and even movement of reagents across membrane surfaces, place the Processor Plus on a level surface. Use Setup 3 and the spirit level to adjust the default level position that the tray assumes when it is not rocking or tilting.

Important When you switch from membrane processing trays to gel staining trays, you must perform the leveling procedure for gel staining trays.

1. Start the leveling procedure with only the tray support on top of the

Processor Plus base.

Outer peristaltic pump tube

Waste tube

Inner peristaltic pump tube



2. Place the spirit level on the flat, clear plate inside the tray support. The level

should sit in a line that is parallel to the direction in which the tray rocks

(Fig 4-3).

Fig 4-3 Placing the level on the clear surface inside the tray support

3. Go to Setup 3. (See page 7 for information on the Setup parameters.)

4. Press" to move the cursor to the second line, then press � or � to adjust the

tray level.)

The angle of the tray changes in response to pressing � or �. The characters on the LCD do not change.

5. When the tray is level, place a blot processing tray on the Processor Plus and

put the lid on the tray.

6. Press" to move the cursor to the first line. Press � to advance to Setup 2 for

and select the tray size that matches the tray you are using.

For Blot processing, use either the Mini or the Standard tray. See page 23 for a description of Setup 2.

7. Press" to move the cursor to the first line. Press � to advance to Setup 7 for

pump calibration.

Level, in a line parallel to the direction of rocking

Setup 3: Adjusttray level up/dn

Setup 3: Adjusttray level up/dn


S e t t i n g U p f o r B l o t P r o c e s s i n g

Calibrating the pump

Use Setup 7 to calibrate reagent delivery. Repeat pump calibration at least once a month and each time you change the tubing in the peristaltic pump. The pump calibration procedure takes five minutes.

Note The calibration procedures for Stain gel and Blot process protocols are different. You must calibrate the pump whenever you switch between these protocol sets.

To calibrate the pump for membrane processing:

1. Place the end of reagent tube 1 in a beaker of at least 400 ml water.

2. Place the end of the drain tube in a 250-ml graduated cylinder or beaker.

3. Select Setup 7 for membranes.

4. Press START.

The message on the screen confirms that you have configured the tubing for calibration.

5. Press START.

The Processor Plus delivers fluid to the tray and then out the drain tube. When the pump stops, measure the amount of water in the graduated cylinder.

6. Press " to move the cursor under the volume values and press � or � to

change the numbers. Enter the amount of water measured in step 5.

7. Replace the end of reagent tube 1 in the beaker of water and the drain tube

in an empty graduated cylinder. Press START.


8. Press START.

The pump again pumps water into the tray and out the drain tube. When the pump stops, measure the amount of water in the graduated cylinder.

Setup 7: CalibPress START

Tube 1 in waterDrn tube in cyl

Measured vol.1= 200 ml

Tube 1 in water Drn tube in cyl




9. Enter the measured volume on the screen and press START.

When calibration is completed, the LCD returns to the protocols.

What’s happening. Processor Plus uses these two values —volume 1 and volume

2— to establish a correlation between the number of times the pump turns and the volume delivered. It then interpolates to determine the number of pump turns necessary to delivery the required volume.


O p e r a t i n g I n s t r u c t i o n s f o r B l o t P r o c e s s i n g

5 Operating Instructions for Blot Processing

To run a blot processing protocol, you must first prepare the tray and reagents.

After you place the tray on the tray support, you must:

• Connect the tubing on the peristaltic pump to the tray lid

• Level the tray (Setup 3)

• Calibrate the pump (Setup 7)

Review the instructions for these procedures on pages 8 –12.

Prepare for blot processing

1. Prepare the membranes for processing. If dry, soak nitrocellulose first in

water and then in buffer. Soak PVDF membranes first in 100% methanol

and then buffer.

Important Keep PVDF membranes immersed in buffer until you are ready to begin processing.

2. Place each membranes in an empty tray chamber.

Place the sides of the membranes that came in contact with the protein facing up.

3. Place the lid with the manifold on top of the disposable tray. Open the valves

on the inlet tube manifold to those tray chambers that contain membranes

(Fig 5-1). Close the valves to those tray chambers that do not contain

membranes.

Fig 5-1 Adjusting valves on the top of the membrane processing tray lid

4. Insert the reagent lines into the proper reagent bottles. See page 54 for

descriptions of pre-programmed blot processing protocols.

The conical centrifuge tubes supplied with the blot processing trays are ideal for holding antibodies. Thread the reagent line through one of the pre-drilled holes in the cap to hold the reagent line firmly while the pump is operating. To assure complete removal of solutions, the tip of the reagent line should extend down into the pointed end of the tube. Do not cover the second hole in the cap, which serves as an air vent.

If you are using other containers for solutions, verify that the tip of the reagent line is submerged in the solution. Tape the reagent line in place at the

Open

Closed


Operating Instructions for Blot Processing

rim of the open container to keep the line secure during pump operation. Do not completely cover the opening, but allow some air to enter to assure accurate delivery.

Note If the Processor Plus is not connected to a serial printer, use Setup 4 to review all volumes. See page 23.

If the Processor Plus is connected to a serial printer, use Setup 6 to verify that the waste bottle will hold all the solutions used in the protocol. See page 24.

Starting at the beginning of the protocol1. From the Main Menu, select Process blot protocols.

2. Press START to go to the first protocol in the set.

3. Press � or � to review the protocols.

Note You can start the Processor Plus at any step in a protocol. To start after step one, see page 15.

4. Press START when the protocol you want appears on the screen.

If you have a serial printer connected to the Processor Plus, the printer begins to print a validation form for the protocol you are about to run.

5. Before the Processor Plus begins the protocol, enter the number of tray

chambers you are using on the top line of the screen. Press � or � to change

the value.

Note The maximum number of chambers for the Mini tray is 4. The maximum number of chambers for the Standard tray is 2.

Verify that the tray description on the second line matches the tray you are using. Press" to move the cursor to the second line on the screen. Press � or � to change the tray size selected.

6. Press START.

As the Processor Plus initiates the protocol, the 10-port valve moves to the port position programmed in the selected first step.

The pump begins to deliver reagent into the level tray and the timer begins counting. When pumping is complete, the tray begins to rock and the screen reports the time remaining in the step and the duration of the entire protocol.

At the end of each step, the timer stops and the tray tilts back as the pump empties the tray.

4 chambersTray: 4 Mini’s

Moving valveto port 5

Step 1, Pumping in from port 5

Step 1 10.0min Total 1:50 hrs

Total time remaining

Time remaining in step


O p e r a t i n g I n s t r u c t i o n s f o r B l o t P r o c e s s i n g

If you have connected a printer to the Processor Plus, at the end of each step the printer records the step number, the length of the step in minutes, the port used and the volume of reagent delivered.

Note The beep option is turned ON in the last step of all preprogrammed protocols.

When the protocol is completed, the Processor Plus emits a series of beeps for 20 seconds if the beep option is ON in the last step. (See page 18.)

7. Run the cleaning protocol, Protocol 5, to clean the reagent lines after

running a protocol. See page 16 for a description of the cleaning protocol.

Starting at a particular step in the protocol

Follow these steps to start a protocol at any step after step 1.

1. Follow steps 1–3 of the procedure on page 14, “Starting at the beginning of

the protocol”.

2. Press EDIT/SET to see the first step in a selected protocol.

Note If you want to edit a step when the instrument is not running, you must press EDIT/SET when finished to get out of the edit mode. See page 17 for more on editing.

3. To review the steps in a protocol, press � or � with the cursor under the step

number.

4. When you find the step at which you want to begin, press START.

Interrupting a protocol

You can temporarily stop or terminate a protocol in progress.

• Press PAUSE to interrupt a protocol without cancelling it.

• Press STOP to interrupt and terminate a protocol in progress.

Pause. You may temporarily stop a protocol to adjust reagent lines and tray

contents.

When you press PAUSE, the timer stops counting and the tray goes to the Level position and stops rocking.

Note If you press PAUSE when the pump is running, the interruption does not begin until the pump is finished.

While the Processor Plus is in PAUSE mode, the tray has three possible positions:

• Level – the tray stops rocking and maintains the level position.

• Recover – the tray tilts forward, in the direction opposite the drain line, so that you can recover antibody solutions for re-use.

• Rock – the tray rocks slowly during the pause.

1. Press � or � to change the tray position during a pause.

2. To end the pause and continue the protocol press START.

STOP. When a protocol is terminated, the screen displays this message.

• Press STOP to return to the protocol list.

Tray — LevelPress Start to continue

Stop — cancel run Start — pump out


Operating Instructions for Blot Processing

• Press START to start the pump and empty fluid in the tray.

You may also manually operate the pump to remove excess reagents from the tray. See “Setup 1: Manual pump operation” on page 22.

Liquid in tray. If you press STOP to cancel the run and return to the protocol list,

liquid in the tray does not get pumped out. The next time you begin a protocol, the Processor Plus displays a reminder.

• Press PAUSE to start the pump and empty fluid in the tray. When the tray is empty, press START to continue.

Cleaning protocol

Note Run Protocol 5 before switching from blot processing to gel staining trays and protocols.

Use blot processing Blot Protocol 5, Clean, to completely rinse the reagent lines and tray with distilled water or TBST (Tris-buffered saline, containing Tween™ 20). The cleaning protocol requires approximately 10 minutes.

On the Mini tray, you may choose to rinse from one to four of the tray chambers. On the Standard tray you may choose to rinse one or two chambers. By default, the tray size corresponds to the last tray you used in a blot processing protocol.

• To change the default tray size, use Setup 2 (see page 23).

1. Place all the reagent lines, except line “0”, into a container with at least

1.5 litres of distilled water or TBST.

2. Place the waste tube from the outer peristaltic pump into the waste bottle.

3. Open the valves on the tray lid to those chambers of the tray you wish to

clean. Close the other valves.

4. Advance to Protocol 5 and press START.

5. Select the number of wells, or tray chambers, you want to clean. If necessary,

move the cursor to the second line on the screen and press � or � to change

the tray size selected.

6. Press START to continue the cleaning protocol.

Liquid in trayPause — pump out

Clean 4 wellsTray: 4 Mini’s


E d i t i n g B l o t P r o c e s s i n g P r o t o c o l s

6 Editing Blot Processing Protocols

There are three types of values you can edit on the Processor Plus: protocol names, step variables, and setup parameters.

Editing a protocol name

A protocol name appears at the beginning of each of the series of 30 steps that make up a protocol. It consists of a number on the first line of the screen and up to 16 alpha-numeric characters on the second line.

The numbers on the first line advance from1 to 10 for the Blot process protocol set. You can change the 16 characters in the second line of any protocol name.

1. Press � or � to review the protocols by number.

When you find the protocol name you want to modify, press ! or " to move the cursor to the second line on the LCD screen.

2. When the cursor is under a character or space you want to modify, press and

hold either � or � to scroll quickly through the available characters.

The available characters include: a blank space, 0–9, A–Z.

3. When finished modifying the protocol name, press ! or " to move the

cursor back to the top line.

Editing step variables

Step variables are parameters that may change in every step of a protocol. All membrane processing protocols contain the following variables:

• step number

• step duration in minutes

• port number for reagent input

• a multiplier for the volume (in ml) of reagent delivered

• end-of-step options: Hold (H), Beep (B) and Rock (R)

Fig 6-1 Variables in each blot processing step

S 1 60.0Min HBRIn5 10ml x2 ^

Step number

Port in

Step duration

H = HoldB = BeepR = Rock

Multiplier

Caret (^) indicates option ON


Editing Blot Processing Protocols

Table 6-1 Step variables and values used in blot processing

Port In

The port in determines the source of the reagent pumped in to the tray. For example, if the Port In is 5, reagent is pumped into the tray from reagent line #5.

Minutes

Minutes are displayed on the LCD in integers and tenths of a minute. The timer begins counting the minutes in each step at the same time that the pump begins pumping reagent into the tray. Depending on the volume of reagent needed, pumping may take up to 1 minute.

MultiplierNote You cannot change the minimum volume assigned to a tray. To deliver less than the minimum volume, fill the reagent bottle with less than the minimum amount.

Use the multiplier to change the volume of reagent delivered to each tray chamber.

The minimum volume is the amount of reagent needed to reliably cover the bottom of a tray chamber. For the Mini tray chambers, it is 10 ml; for the Standard tray chambers, it is 25 ml. These minimum volumes are stored in the Processor Plus memory and cannot be modified. You can increase the volume delivered to a chamber by selecting a multiplier that is greater than 1.

In each protocol step, the Processor Plus determines the total volume to be pumped by calculating the product of the minimum tray volume, the multiplier, and the number of tray chambers being used.

Note The minimum volume is adequate at room temperature. When running the Processor Plus in a cold room or overnight, you may have to increase the multiplier to ensure sufficient reagent.

For example, in Figure 6-1 on page 17 the multiplier in Step 1 is 2. If 3 chambers were selected at the beginning of the protocol, the pump will deliver 60 ml (10 ml × 2 × 3) of the reagent at port 5.

Hold, Beep and Rock (HBR)

Hold, Beep and Rock—H, B, and R—are options that can be turned off or on. Table 6-2 on page 19 shows the effect of having different combinations of these options on at the end of a step.

To turn an option off or on, move the cursor below the H, B, or R, then press � or �. A caret (^) indicates the option is turned on.

Note You must set Hold to on if you want reagent to stay in the tray at the end of the step. If only Beep and/or Rock are on, reagent is pumped out of the tray at the end of the step.

Variable Available values

Steps 1–30

Port In* 1–9

Minutes 0.1–900.0

Multiplier 1–5

Hold on (^) or off

Beep on (^) or off

Rock on (^) or off

* Port 0 is reserved as an air vent in blot processing protocols



Table 6-2 Possible H, B, R combination outcomes

Editing when the Processor Plus is not running a protocol

Edit when the Processor Plus is not running a protocol to save the changes in memory.

1. Select the protocol you want to edit.

2. Press EDIT/SET to see the first step in the protocol.

3. Press � or � to review each step in the protocol. You can edit the variables at

each step. Table 6-1 on page 18 shows the values available for each variable.

• Press ! or " to move the cursor from one variable to the next. To switch H, B, or R, first move the cursor to the bottom, right corner of the screen.

• Press � or � to change the value of the selected variable. When a caret (^) appears below H, B, or R, it indicates that option is ON.

4. Press EDIT/SET to complete the edit, save changes and return to the

beginning of the protocol.

Use Setup 5 to print a record of the revised protocol. See page 23.

Adding or inserting a step

You can add a step after any step in a protocol. The variables in the added step contains the same values as the step that precedes it. Edit the variables in the new step as needed.

1. Follow steps 1 through 3 of the previous editing procedure to find the place

in the protocol to add a step.

2. Press ADD to insert a new step after the step that is visible on the screen. The

new step has the next step number and the variable values found in the step

that precedes it. The step number of all subsequent steps increases by one.

H B R Outcome

^ Protocol pauses at end of step and reagent remains in level tray until you press START to continue.

^ ^ Protocol pauses at end of step, reagent remains in level tray and beep sounds every six seconds until you press START to continue. Without attention, the beep stops after five minutes.

^ ^ Protocol pauses at end of step, reagent remains in rocking tray until you press START to continue.

^ ^ ^ Protocol pauses at end of step, beep sounds every six seconds, reagent remains in rocking tray until you press START to continue. Without attention, the beep stops after five minutes.

^ Beep sounds at end of step.


Editing Blot Processing Protocols

Deleting a step

You can delete any step in a protocol. After you delete a step, the screen displays the step that preceded the deleted step. The step number of all subsequent steps decreases by one.

1. Follow steps 1 through 3 of the procedure for editing (page 19) to find the

step you want to delete.

2. Press DEL to delete the step that is visible on the screen.

Editing a protocol in progress

You can edit the protocol in progress while the Processor Plus is running. These protocol changes are not permanent; the protocol in memory remains unchanged. To save protocol changes, you must edit the protocol when it is not in progress.

You can make the following protocol edits when the Processor Plus is running:

• Change the time only in the current step

• Change the time, port number, multiplier, or switches in subsequent steps

• Add or delete steps after the current step

Important You cannot interrupt pump operation. Do not press EDIT/SET while the pump is running.

Note In the current step, you can only change time. You must press EDIT/SET to change variables in subsequent steps.

To change the time in the current step

• Move the cursor to the time and press � or � to change the value.

To change a value in a subsequent step

1. Press EDIT/SET to go into edit mode.

The LCD displays the next step in the protocol.

2. Move the cursor under Step number, advance to the step you want to edit

and edit the values in that step.

3. When you finish editing, press EDIT/SET to return to the step in progress.

To add one or more steps

Note You cannot add more steps during the last step.



2. Move the cursor under Step number and advance to the step that precedes

the point where you want to add steps.

3. Press ADD once for each step you want to add. Edit the new steps as needed.




To delete one or more steps



2. Move the cursor under Step number and advance to the step you want to

delete.

3. Press DEL once for each step you want to delete.


Note If you delete the step that follows the step in progress, the software goes out of edit mode and returns to the step in progress.

Power outages. If a power outage occurs after you have edited the protocol in

progress, the changes you made to the protocol are lost.

If a power outage occurs while you are editing a protocol in progress, the instrument displays the Main menu when power returns. Restart the protocol at the interrupted step. See page 15 for directions.


Setup Parameters for Blot Processing

7 Setup Parameters for Blot Processing

Setup parameters are values that apply to an entire set of protocols. You cannot use the setup parameters when a protocol is running.

The Processor Plus has seven Setup parameters for blot processing:

1. Manual pump operation

2. Tray size

3. Tray level

4. Display protocol volume

5. Print protocol steps

6. Print total volumes of reagents used in the protocol

7. Pump calibration

To go to Setup menu for blot processing:

Note Be sure to begin from Process blot to edit Setup parameters for blot processing

• In any process blot protocol, press and hold EDIT/SET until the screen displays the Setup parameters.

Setup 1: Manual pump operation

Use to manually operate the pump when a protocol is not running. In Setup 1, the pump by default operates as long as is necessary to deliver the same volume it delivered in the last completed step.

1. Go to Setup 1 for blot processing.

2. Press" to move the cursor under the port number, then press � or � to

select a port.

3. Press" to move the cursor to the second line. Press � or � to switch between

in and out.

• Use in to pump fluid into the tray through a port

Note In blot processing Setup 1, with the tray lid in place, pumping out pumps some air from the tray lid through the selected port.

• Use out to pump fluid out of the tray and into a reagent or waste bottle.

Press START to manually start the pump.

4. When the pump stops, press" to move the cursor back under the Setup

number, then press � or � to review the other Setup parameters or press

STOP to return to protocols.

Setup 1: Port 3 pump out


S e t u p P a r a m e t e r s f o r B l o t P r o c e s s i n g

Setup 2: Tray size

Use to choose the tray size. Each protocol set has two pre-defined tray sizes.

Note The minimum volume is adequate at room temperature. When running the Processor Plus in a cold room or overnight, you may have to increase the multiplier to ensure sufficient reagent.

A minimum volume in millilitres is assigned to each tray size, based on the capacity of the tray (Table 7-1). You cannot edit tray sizes or the minimum volume.

Table 7-1 Minimum Volume for Tray Sizes

To use Setup 2:

1. Select Setup 2 for Process blot.

2. Press" to move the cursor to the second line. Then press � or � to select a

tray.

3. Press" to move the cursor to the top line. Press � or � to go to another

setup parameter or press STOP to return to protocols.

Setup 3: Tray level

Use to adjust the level position of the tray, when it is not rocking or tilting. See page 9 for instructions on leveling the blot processing trays.

Setup 4: Display protocol volume

Use to display the total reagent volume needed for each port for any blot processing protocol.

Note Setup 4 only displays the volumes on the LCD. Use Setup 6 to print a report of the total reagent volume needed at each port.



protocol.

3. Press START.

On the top line, enter the number of chambers. Press" to move the cursor to the second line and select the appropriate tray size.

Blot Process Tray Minimum Volume

Mini 10 ml

Standard 25 ml

Setup 2: Tray: 4 Mini

Setup 4: DisplayVol of Prot. 2



Setup Parameters for Blot Processing

4. Press START.

The LCD displays the total volume of reagent needed at Port 1 for the protocol, number of chambers and tray size specified.

5. Continue to press START to see the volumes needed at each port.

Setup 5: Print protocolNote See page 4 for directions on connecting the Processor Plus to a serial printer.

Use to print out the steps in a protocol when a protocol is not running.



protocol.

3. Press START.

The serial printer produces a record of all the step variables in the selected protocol.

Setup 6: Print volume of protocolNote See page 4 for directions on connecting the Processor Plus to a serial printer.

Use Setup 6 to print a report of total reagent volume needed for each port and the total of all volumes.

Note The value of the total volumes varies, depending on the tray size selected and the number of chambers used.



protocol.

3. Press START.

On the top line, enter the number of chambers. Press" to move the cursor to the second line and select the appropriate tray size.

Port 1 totalvolume = 40ml

Setup 5: PrintProtocol # 10

PrintingProtocol # 10

An example of a volume report printout

Setup 6: PrintVol of Prot. 8



S e t u p P a r a m e t e r s f o r B l o t P r o c e s s i n g

4. Press START.

The serial printer generates a report of the total volumes needed at each port for the selected protocol, based on the number of chambers used, the tray size and the multiplier in each step.

Setup 7: Pump calibration

Use Setup 7 to calibrate volume delivery. Calibrate the pump at least once a month and each time you change the tubing in the peristaltic pump.

Note The pump calibration for blot processing is not the same as pump calibration for gel staining. You must calibrate the pump when you switch between protocol sets.

See page 11 for pump calibration instructions.

PrintingVol of Prot. 8


Setting Up for Gel Staining

8 Setting Up for Gel Staining

The Hoefer Processor Plus allocates memory for fourteen gel staining protocols; nine of these protocols are pre-programmed (Table 8-1). Each protocol can contain up to 30 steps. See page 33 for directions on editing these protocols.

The Processor Plus has two gel staining trays (Table 8-2) and each tray has its own tray base and lid.

Table 8-1 Pre-programmed gel staining protocols

Table 8-2 Tray sizes for gel staining

Important Thoroughly rinse the staining tray with deionized water after each use. Clean and dry instrument surfaces after each use. To avoid scratching the PTFE coating, do not use metallic instruments on this surface.

Important Do not introduce any hot volatile reagents into this instrument.

Protocol Name Applications

1 DNA silver DNA Gels 1.0 mm unbacked gels0.5 mm backed gels

2 Protein silver SDS and Native 0.75 and 1.0 mm unbacked gels0.5 mm backed gelsImmobiline DryPlates

3 Protein silver SDS and Native 1.5 mm unbacked gels

4 Protein silver IEF 0.5 and 1.0 mm unbacked gels

5 Protein Coomassie SDS and Native 1.0 mm unbacked gels0.5 mm backed gels

6 Protein Coomassie SDS and Native 0.75 mm unbacked gels

7 Protein Coomassie SDS and Native 1.5 mm unbacked gels

8 Protein Coomassie IEF 0.5 and 1.0 mm unbacked gels

9 Clean Rinses reagents out of tubings, valves and ports

Tray Number of gels

Gel Size (cm)

Reagent Capacity(ml/chamber)

19 × 29 cm

29 × 35 cm

1 12 4

1 2 22 6

14 × 16 11 × 2611 × 13 7 × 8

20 × 26 14 × 1611 × 13 11 × 26 7 × 8

125 – 200

250 – 400


S e t t i n g U p f o r G e l S t a i n i n g

Configuring the gel staining tray and pump tubing assembly

For gel staining, only one tube connects the gel staining tray to the inner peristaltic pump (Fig 8-1). The tray lid has no tubing inlets. Gel staining protocols do not use the outer peristaltic pump tube.

When the appropriate male and female Quick-Fit couplings are pressed together, you can hear a click as they connect.

Fig 8-1 Gel staining tube and tray configuration

Leveling the tray

For reliable and even movement of reagents across gels, place the Processor Plus on a level surface. Use Setup 2 to select the tray size that matches the tray you are using. Use Setup 4 and the spirit level to adjust the default level position that the tray assumes when it is not rocking or tilting.

Important Each tray has its own level position. Each time you change trays, use the spirit level to verify that the tray is level.

1. Place a gel staining tray on the Processor Plus and put the lid on the tray.

2. Go to Setup 2 and select the tray size that matches the tray you are using.

For gel staining, use either 19 × 29 cm or 29 × 35 cm. (See page 39 for a description of Setup 2.)

3. Place the level on the tray lid. The level should sit in a line that is parallel to

the direction in which the tray rocks (Figure 8-2 on page 28).

Tray drain tube

Inner peristaltic pump tube

Outer peristaltic pump tube


Setting Up for Gel Staining

Fig 8-2 Placing the level on the surface of the tray lid, viewed from above

4. From Setup 4 for Stain gel, press" to move the cursor to the second line,

then press � or � to adjust the tray level up or down.

The angle of the tray changes in response to pressing � or �. The characters on the LCD do not change.

5. When the tray is level, press" to move the cursor to the first line. Press � to

advance to Setup 6, pump calibration.

Calibrating the pump

Use Setup 6 to calibrate reagent delivery. Repeat pump calibration at least once a month and each time you change the tubing in the peristaltic pump.

Note The calibration procedures for Stain gel and Blot process protocols are different. You must calibrate the pump whenever you switch between the two protocol sets.

To calibrate the pump for gel staining:

1. Place the end of reagent tube 1 in a beaker of at least 400 ml water.

2. Place the end of tube 0 in a 250-ml graduated cylinder or beaker.

3. Select Setup 6 for membranes.

4. Press START.


5. Press START.

Level

Direction of tray rocking

Fron

t of t

ray

Setup 4: Adjust tray level up/dn

Setup 6: CalibPress START

Tube 1 in water Tube 0 in cyl


S e t t i n g U p f o r G e l S t a i n i n g

The Processor Plus delivers fluid to the tray and then out tube 0. When the pump stops, measure the amount of water in the graduated cylinder.

6. Press " to move the cursor under the volume values and press � or � to

change the numbers. Enter the amount of water measured in step 5.

7. Replace the end of reagent tube 1 in the beaker of water and tube 0 in an

empty graduated cylinder or beaker. Press START.


8. Press START.

The pump again pumps water into the tray and out tube 0. When the pump stops, measure the amount of water in the graduated cylinder.

9. Enter the measured volume on the screen and press START.

When calibration is completed, the LCD returns to the protocols.

What’s happening. Processor Plus uses these two values —volume 1 and volume

2— to establish a correlation between the number of times the pump turns and the volume delivered. It then interpolates to determine the number of pump turns necessary to delivery the required volume.


Tube 1 in water Tube 0 in cyl


Calibrationcomplete


Operating Instructions for Gel Staining

9 Operating Instructions for Gel Staining

To run a protocol, you must first prepare the tray and reagents.

The first time you prepare the tray, you must:

• Configure the tubing on the peristaltic pump

Note Each time you change tray sizes, select the tray size in Setup 2 and level the tray in Setup 4.

• Level the tray (Setup 4)

• Calibrate the pump (Setup 6)

Review the instructions for these procedures on pages 27 – 29.

Prepare for gel staining Note Calibrate the pump whenever you change tubing in the peristaltic pump, or at least once a month.

1. Carefully place the gels in the empty tray.

Place plastic-backed gels with the backing side down. If using gels without backing, you may trim off any low-percentage stacking layer before staining.

2. Place the magnets on the tray as needed to prevent gels from overlapping or

piling up around the drain.

Use the magnets to create separate processing areas for multiple gels. The magnets can also be placed on the corners of plastic-backed gels to keep them submerged.

3. Place the plain glass lid on the tray. Note the moulded indentations at both

sides allow easy lid placement and removal.

4. Put the ends of the reagent lines into the appropriate reagent bottles.

Refer to the Automated Gel Stainer Protocol Guide for descriptions of each gel staining protocol. See also “Pre-programmed Staining Protocols” on page 58.

Starting at the beginning of the protocolNote You can start the Processor Plus at any step in a protocol. To start after step one, see page 31.

1. From the Main Menu, select Stain gel protocols.

2. Press START to go to the first protocol in the selected protocol set.

3. Press � or � to review the protocols in the selected protocol set.

4. Press START when the protocol you want appears on the screen.

If you have a serial printer connected to the Processor Plus, the printer begins to print a validation form for the protocol you are about to run.

5. On the top line, verify the volume of reagent to be pumped for each step.

Press � or � under a number to change its value.

6. Press" to move the cursor to the second line and verify the tray selection.

Press � or � to select the appropriate tray description, then press START.

Volume: 175 ml Tray: 19 x 29 cm


O p e r a t i n g I n s t r u c t i o n s f o r G e l S t a i n i n g

As the Processor Plus initiates the protocol, the 10-port valve moves to the port position programmed in the selected first step. The pump begins to deliver reagent into the tray, the tray rocks and the timer begins counting.

Note The tray rocks during every step.

When pumping is complete, the screen reports the time remaining in the step and the duration of the entire protocol.

At the end of each step, the timer stops and the tray tilts back as the pump empties the tray.

If you have connected a printer to the Processor Plus, at the end of each step the printer records the step number, the length of the step in minutes and the ports used.

When the protocol is completed, the Processor Plus emits a series of beeps for 20 seconds if the beep option is ON in the last step. (See page 34.)

Important Clean the staining tray thoroughly after each use. See page 47 for cleaning instructions. See page 32 for a description of Protocol 9, the cleaning protocol.

Starting at a particular step in the protocol

Follow these steps to start a protocol at any step after step 1.

1. Follow steps 1–3 of the previous procedure on page 30.

2. Press EDIT/SET to see the first step in the protocol.

Note If you edit a step when the instrument is not running, you must press EDIT/SET when finished to end edit mode. See page 33 for more on editing.

3. To review the steps in a protocol, press � or � with the cursor under the step

number.

4. When you see the step at which you want to begin, press START.

Interrupting a protocol

You can temporarily stop or terminate a protocol in progress.

• Press PAUSE to interrupt a protocol without cancelling it.

• Press STOP to interrupt and terminate a protocol in progress.

Pause. You may temporarily stop a protocol to adjust reagent lines and tray

contents. See page 20 for directions on editing a protocol in progress.

Moving valveto port 1

Step 1, Pumping in from port 1

Step 1 9.3 minTotal 0:10 hrs

Time left in step

Total time left in protocol


Operating Instructions for Gel Staining

Note If you press PAUSE when the pump is running, the interruption does not begin until the pump is finished

When you press PAUSE, the timer stops counting and the tray goes to the Level position and stops rocking.

While the Processor Plus is in PAUSE mode, the tray has three possible positions:

• Level – the tray stops rocking and maintains the level position defined in Setup 4.

• Recover – the tray tilts forward so that you can recover solutions.

• Rock – the tray rocks slowly during the pause.

1. Press � or � to change the tray position during a pause.

2. To continue the protocol after a pause, press START.

STOP. When the protocol is terminated, the screen displays this message.

• Press STOP to return to the protocol list.

• Press START to start the pump and empty the tray.

You may also manually operate the pump to remove excess reagents from the tray. See page 38.

Cleaning protocol

Note Run Protocol 9 before switching from gel staining to blot processing trays and protocols.

Use gel staining Gel Protocol 9, Clean, to completely rinse the tubing and tray with distilled water. The cleaning protocol requires approximately 10 minutes.

1. Place the reagent line labelled “0” into a container with at least 1.5 litres of

distilled or deionized water. If you are using the 29 × 35 tray, the container

should hold 3 litres of distilled or deionized water.

2. Place all the other reagent lines into a waste bottle that can hold at least

4 litres.

3. Advance to Protocol 9 and press START.

4. Adjust the volume and tray size, if necessary, and press START.

Tray — LevelPress Start to continue

Stop — cancel run Start — pump out

Volume: 175 ml Tray: 19 x 29 cm


E d i t i n g G e l S t a i n i n g P r o t o c o l s

10 Editing Gel Staining Protocols

Important Design gel staining protocols so that the same reagent lines always deliver the same reagents. This consistency minimizes contamination of the reagent lines and decreases the potential for mixing incompatible reagents.

There are three types of values you can edit on the Processor Plus: protocol names, step variables, and setup parameters.

Editing a protocol name

A protocol name appears at the beginning of each of the series of steps that make up a protocol. It consists of a number on the first line of the screen and up to 16 alpha-numeric characters on the second line. The numbers on the first line advance from 1 through 14 for gel staining protocols. You can change any of the 16 characters in the second line of the protocol name.

1. From the Main Menu, select the Stain gel protocols and press START.

2. Press � or � to review the protocols by number.

When you see the protocol name you want to modify, press ! or " to move the cursor to the second line on the LCD screen.

3. When the cursor is under a character or space you want to modify, press and

hold either � or � to scroll quickly through the available characters.

The available characters include: a blank space, 0–9, A–Z.

4. Press ! or " to move the cursor back to the first line.

Editing step variables

Step variables are values that may change in every step of each protocol (Fig 10-1). The LCD screen displays each step in a protocol. Steps in all gel staining protocols contain the following variables:

• step number

• port number for reagent input

• step duration in minutes

• end-of-step options: hold (H), beep (B) and rock (R)

• port number for reagent output

Fig 10-1 Variables in each gel staining step

S 1 60.0 min HBRIn5 out 7 ^

Step number

Step duration

H, B, Roptions

Port in Port out

Caret (^) indicates option ON


Editing Gel Staining Protocols

Table 10-1 Step variables and values used in gel staining protocols

Port In

The port in defines the reagent line through which reagent enters the tray. For example, if the Port In is 5, reagent is pumped into the tray from reagent line #5.

Minutes

Minutes are displayed on the LCD in integers and tenths of a minute. The timer begins counting the minutes in each step at the same time that the pump begins pumping reagent into the tray. Depending on the volume of reagent needed, pumping may take up to 1 minute.

Port Out

Port out defines the reagent line used to carry fluid out of the tray. You can collect all waste in one bottle or divide it according to composition. For example, the pre-programmed protocol for silver staining divides waste into three bottles:

• waste that contains silver into one waste bottle, via Port Out 9

• waste that contains formaldehyde or glutardialdehyde into the second waste bottle, via Port Out 8

• waste that contains waste water, ethanol, acetic acid, EDTA and glycerol into a third waste bottle, via Port Out 7.

Hold, Beep and Rock

Hold, beep, and rock are options that can be turned off or on. Table 10-2 on page 35 shows the effect of having different combinations of these options on in a step.

To turn an option off or on, move the cursor below the H, B, or R, then press � or �. A caret (^) indicates the option is on.

Note You must switch Hold to on if you want reagents to stay in the tray at the end of the step. If only Beep and/or Rock are on, reagent is pumped out of the tray at the end of the step.

Variable Available values

Steps 1–30

Port In 0–9

Minutes 0.1–900.0

Port out 0–9

Hold on (^) or off

Beep on (^) or off

Rock on (^) or off



Table 10-2 Possible HBR combination outcomes

Editing when the Processor Plus is not running a protocol

Edit when the Processor Plus is not running a protocol to save the changes in memory.

1. From the Main menu, select Stain gel protocols. Press START to go to the

first protocol.

2. Press � or � to review the protocols in the protocol set.

3. Press EDIT/SET to see the first step in a protocol.

If necessary, press ! or " to move the cursor under the step number.

4. Press � or � to advance the step number through a protocol. You can edit

the variables at each step. Table 10-1 on page 34 shows the values available

for each variable.

• Press ! or " to move the cursor from one variable to the next. To change the status of H, B, or R, move the cursor to the bottom, right corner of the screen.

• Press � or � to change the value of the selected variable. When a caret (^) appears below H, B, or R, it indicates that option is ON.

5. Press EDIT/SET to complete the edit, save the changes and return to the

protocols.

Use Setup 5 to print a record of the revised protocol. See page 40.

H B R Outcome

^ Protocol pauses at end of step and reagent remains in level tray until you press START to continue.

^ ^ Protocol pauses at end of step, reagent remains in level tray and beep sounds every six seconds until you press START to continue.

^ ^ Protocol pauses at end of step, reagent remains in rocking tray until you press START to continue.

^ ^ ^ Protocol pauses at end of step, beep sounds every six seconds, reagent remains in rocking tray until you press START to continue.

^ Beep sounds at end of step.


Editing Gel Staining Protocols

Adding or inserting a step

You can add a step after any step in a protocol. The variables in the added step contains the same values as the step that precedes it. Edit the variables in the new step as needed.

1. Follow steps 1 through 3 of the previous editing procedure, (page 35) to find

the place in the protocol where you want to add a step.

2. Press ADD to insert a new step after the step on the screen. The new step has

the next step number but contains the variable values found in the step that

precedes it. The step number of all subsequent steps increases by one.

Note The Processor Plus contains empty protocols that contain only one step. Add steps to these protocols to enter your own custom protocols.

Deleting a step

You can delete any step in a protocol. After you delete a step, the screen displays the step that preceded the deleted step. The step number of all subsequent steps decreases by one.

1. Follow steps 1 through 3 of the editing procedure (page 35) to find the step

you want to delete.

2. Press DEL to delete the step on the screen.

Editing a protocol in progress

You can edit the protocol in progress while the Processor Plus is running. These protocol changes are not permanent; when the protocol ends it reverts to its original definition. To save protocol changes, you must edit the protocol when a it is not in progress.

You can make the following protocol edits when the Processor Plus is running:

• Change the time only in the current step

• Change the time, port in, port out, or end-of-step options in subsequent steps

• Add or delete steps after the current step

Important You cannot interrupt pump operation. Do not press EDIT/SET while the pump is running.

Note In the current step, you can only change time. You must press EDIT/SET to change variables in subsequent steps.

To change the time in the current step

• Move the cursor to the time and press � or � to change the value.

To change a value in a subsequent step



2. Move the cursor under Step number, advance to the step you want to edit

and edit the values in that step.




To add one or more steps



2. Move the cursor under Step number and advance to the step that precedes

the point where you want to add steps.

3. Press ADD once for each step you want to add. Edit the new steps as needed.


Note During the last step, you cannot add more steps.

To delete one or more steps



2. Move the cursor under Step number and advance to the step you want to

delete.

3. Press DEL once for each step you want to delete.


Note If you delete the step that follows the step in progress, the software goes out of edit mode and returns to the step in progress.

Power outages. If a power outage occurs after you have edited the protocol in

progress, the changes you made to the protocol are lost.

If a power outage occurs when you are in edit mode and a protocol is in progress, the instrument does not return to the protocol when power returns. Instead it displays the Main menu. To continue the interrupted protocol, restart at the interrupted step. See page 31 for directions.


Gel Staining Setup Parameters

11 Gel Staining Setup Parameters

Setup parameters are values that apply to an entire set of protocols. You cannot use the setup parameters when a protocol is running.

The Processor Plus has six Setup parameters for gel processing:

1. Manual pump operation

2. Tray size

3. Volume delivered

4. Tray level

5. Print protocol

6. Pump calibration

To go to the Setup mode for gel staining:

1. In the Main Menu, select Stain gel.

Note Be sure to begin in Stain gel to see the Setup parameters for gel staining.

2. Once you are in the gel staining protocol set, press and hold EDIT/SET until

the screen displays the Setup parameters.

Setup 1: Manual pump operation

Use Setup 1 to manually operate the pump when a protocol is not running. In Setup 1, the pump by default operates as long as is necessary to deliver the same volume it delivered in the last step completed.

1. From Setup 1 for Stain gel, press ! or " to move the cursor under the port

number, then press � or � to select a port.

2. Press ! or " to move the cursor to the second line, under in or out. Press �

or � to switch between in and out.

• Use in to pump fluid into the tray through a port

• Use out to pump fluid out of the tray, into a reagent or waste container

Press START to manually start the pump.

3. When the pump stops, press" to move the cursor back under the Setup

number, then press � or � to review the other Setup parameters, or press

STOP to return to protocols.

Setup 1: Port 3 pump out


G e l S t a i n i n g S e t u p P a r a m e t e r s

Setup 2: Tray size

Use to review the available tray sizes. Gel stain has two pre-defined tray sizes.

Note You can also modify the tray size and volume at the beginning of a protocol.

You cannot edit tray sizes, but you can edit the volume associated with the tray in Setup 3. Use the Setup 2, tray size, in conjunction with Setup 3, Volume Delivery. Each tray size in Setup 2 determines the range of available values in Setup 3. See Table 11-1.

Table 11-1 Relation of tray sizes to values in Setup 3

1. From Setup 2 for Stain gel, press " to move the cursor to the second line.

Then press � or � to review the tray options.

2. Select a tray size.

3. Press " to move the cursor to the top line and then press � to advance to

Setup 3: Volume Delivery.

Setup 3: Volume delivery

Use to define the of reagent (ml) delivered to the tray in each step of the gel staining protocols.

1. From Setup 3 for Stain gel, press" to move the cursor to the second line,

under the number you want to modify. Then press and hold � or � to

change the number.

2. When finished, press" to move the cursor to the first line. Press � or � to

advance to another Setup parameter or press STOP to return to protocols.

Setup 4: Tray level

Use to adjust the level position of the tray, when it is not rocking or tilting. See page 27 for directions on using Setup 4.

Setup 2 Tray Setup 3 Range(ml)

Recommended Volume (ml)

19 × 29 cm 125–200 125

29 × 35 cm 250–400 250

Setup 2: Tray19 x 29 cm

Setup 3 Volumepumped = 175 ml


Gel Staining Setup Parameters

Setup 5: Print protocolNote See page 4 for directions on connecting the Processor Plus to a serial printer.

Use to print out the steps in a protocol either before or after running a protocol.

1. Select Setup 5 for Gel stain.


protocol.

3. Press START.

Setup 6: Pump calibration

Use to calibrate reagent delivery. Repeat calibration at least once a month and each time you change the tubing in the peristaltic pump.

Important The calibration procedures for Stain gel and Process blot protocols are different. You must calibrate the pump for each set of protocols.

To calibrate the pump for gel staining, see page 28.

Setup 5: PrintProtocol # 14

PrintingProtocol # 14


U s i n g t h e p r o t o c o l k e y

12 Using the protocol key

Note The Processor Plus cannot read the protocol key when it is in the Edit mode or in one of the Setup menus.

When you insert the protocol key into the protocol key slot, the instrument reads the key and writes it in the last protocol in the protocol set. Any protocol that is already in memory in this last position will be overwritten by the protocol on the protocol key.

• If the protocol on the protocol key is a gel staining protocol, it is written into Gel Protocol 14. You can copy a gel staining protocol from the key into any of the 14 gel staining protocols on the Processor Plus.

• If the protocol on the protocol key is a blot processing protocol, it is written into Blot Protocol 10. You can copy a blot processing protocol from the key into any of the 10 blot processing protocols on the Processor Plus.

You can also copy any protocol from the Processor Plus onto the protocol key.

Note You cannot copy a gel staining protocol on the key into memory on the Processor Plus that is reserved for a blot processing protocol, or vice versa.

Editing with the protocol key in the slot. If you leave the protocol key in the

slot while you edit the last protocol in the protocol set—Protocol 10 in blot processing or Protocol 14 in gel staining— the Processor Plus automatically overwrites the protocol on the key when you press either STOP or EDIT/SET to exit the edit mode.

Important To prevent accidentally overwriting the protocol on the protocol key, remove the protocol key before you begin editing protocols.

To copy a protocol from the key to the Processor Plus

Note If you copy a protocol from the key into a protocol on the Processor Plus that already contains defined steps, you overwrite those steps and replace them with the protocol on the key.

1. With the key in the protocol key slot, use the arrow keys to advance to a

protocol number on the Processor Plus.

2. Press DEL.

The Processor Plus writes a copy of the protocol on the key into the selected protocol number. Remove the protocol key to prevent accidentally overwriting the protocol on the key.

Reading protocol key...Please wait

Protocol 14MyNew Stain

Reading protocol key...Please wait


Using the protocol key

To copy from the Processor Plus to the protocol key

The protocol key can hold only one protocol at a time. When you copy a protocol onto the key, you overwrite the protocol already on the key.

1. With the key in the protocol key slot, use the arrow keys to advance to the

protocol number on the Processor Plus that you want to copy onto the key.

2. Press ADD.

The Processor Plus emits one long beep as it copies the protocol onto the key.

To edit a protocol on the protocol key

To edit the protocol on the protocol key, copy an edited protocol from the Processor Plus on to the protocol key.

Writing protocol key...Please wait


T r o u b l e s h o o t i n g

13 Troubleshooting

Pumping irregularities. If you observe a pumping irregularity while a protocol is

in progress, record the port currently indicated on the LCD. Then press STOP to terminate the protocol. Remove the gels or membranes from the tray. Remove the reagent line from the reagent bottle and insert it into a bottle of deionized water. Pump water in and out several times to dislodge any precipitates or obstructions. If unable to clear the reagent line, replace it with a new line of the same length and diameter.

Power Outage. If a power outage of 10 minutes or less occurs during a protocol,

the Processor Plus resumes the interrupted step when power returns. See page 21 and page 37 for additional information about power outage consequences.

Instrument-based problems

Symptom Possible cause Recommended action

Diagnostics cycle indicates component failure.

Component failure. Press START to continue to next test. Call your local Amersham Biosciences office for further information.

Pumped volume higher than expected volume.

Tray size being used does not match tray size in Setup 2.

Check the tray size selected in Setup 2.

Incorrect reagent line assemblies.

Use only reagent lines supplied with the Processor Plus. Do not shorten the tubing. Do not attach pipettes in place of rigid, opaque tubing.

Pump out of calibration. Calibrate the pump.

Pumped volume lower than expected volume.

Tray size being used does not match tray size in Setup 2.


Incorrect reagent line assemblies.

Use only the reagent lines supplied with the Processor Plus.

Reagent line has come out of reagent bottle or the liquid level is below the end of the opaque rigid tubing.

Make sure rigid tubings are fully immersed in the reagent bottle and there is sufficient reagent in the bottle.

Reagent tubing damaged or cracked.

Replace reagent tubing.

Obstructions or constriction in reagent line or 10-port valve.

Run the cleaning protocol or replace the reagent line.

Pump head tubing worn. Replace the tubing and recalibrate the pump.

Leaks in reagent line at staining tray drain port or pump head tubing inlet or outlet.

Reagent line connection loose or improperly attached.

Adjust fitting of reagent line and connection.

Leaks in reagent lines at 10-port valve.

Positioning controls out of adjustment.

Turn off mains power switch and then turn on again to test the positioning controls.


Troubleshooting

Instrument-based problems, continued


Pump does not empty the tray completely.

Obstruction in splash guard. Clean the splash guard (staining tray).

Tray not level. Adjust the tray level.

Obstructions or constriction in reagent lines.

Run the cleaning protocol to clear obstructions or change the reagent lines.

Solution is splashing against the lid.

Tray or unit not level. Make sure unit sitting on level surface. Adjust the tray level.

Tray used does not match tray selected in Setup 2.


Liquid accumulating under pump.

Pump tubing worn. Replace the pump tubing.

Gel Staining problemsGel sticks to the staining tray.

Insufficient reagent volume in Setup 3.

Increase the volume delivered in Gel stain Setup 3.

Check the troubleshooting section of the Protocol Guide for modifications that may help prevent gel sticking.

Uneven staining. Reagent buildup on tray and reagent lines.

Rinse the staining tray with distilled water after each use.

Gels sticking to tray surface. Check the troubleshooting section of the Protocol Guide for modifications that may help prevent gel sticking.

Gels crowded or overlapping in tray.

Make sure each gel has adequate processing area. Rearrange the magnets to allow more movement.

Contaminants in reagent lines. Always run the cleaning protocol before switching to a different protocol to minimize mixing of reagents.

Insufficient reagent volume in Setup 3.

Increase the delivered volume to ensure that the gel is fully submerged. It may help to turn the gels over during the staining process to increase exposure to reagents.

Check the troubleshooting section of the Protocol Guide for more information.

High background or poor sensitivity

Check the troubleshooting section of the Protocol Guide for more information.


T r o u b l e s h o o t i n g

Blot Processing problems


Weak or no signal. Inefficient transfer. Confirm protein transfer by staining residual gel.

Check nitrocelullose or PVDF membranes for sample with Ponceau S reversible stain. With nylon, use Rainbow Molecular Weight Markers instead of Ponceau S.

Confirm that the electrical conditions and transfer buffers used match the recommendations of the manufacturer of the transfer unit.

Transfer of higher mass proteins can be improved by extending the transfer time or by using a lower percentage separating gel.

Check for uniform, bubble-free contact between the gel and membrane during transfer.

Poor binding of proteins to membrane.

Do not used aged membranes, which may not wet properly or may not bind proteins.

Add methanol in Towbin buffer to improve the protein binding to membranes.

Add 0.1% SDS to improve transfer from the gel. (SDS may impair binding efficiency to nitrocellulose membranes.)

Gel transferred the wrong direction.

Use Rainbow Molecular Weight Markers to confirm transfer or stain membrane with Ponceau S prior to processing. Do not use Ponceau S with nylon.

Insufficient protein loaded on the gel.

Load more protein.

Antibody concentration too low. If commercially prepared, follow manufacturers suggested dilutions, titre up and down from the starting point. Concentration and titre of antibodies are both important.

Some antigens may be affected by the treatments required in electrophoresis.

Test binding efficiency with slot blot or other blotting method. Denaturation during SDS electrophoresis may affect antibody binding.

Detection reagents degraded or not properly stored.

Prepare fresh reagents.

Membrane degraded. Keep nitrocellulose in a closed bag away from heat, light and moisture. It will degrade if not stored properly. Check membrane wetting. Nitrocellulose should quickly absorb water. PVDF should be wet in methanol prior to equilibration in transfer buffer.


Troubleshooting

Blot Processing problems, continued


Weak or no signal. Antibodies/reagents not delivered.

Check that the correct reagent line is in the vessel and that the opaque reagent tubing reaches to the bottom of the container.

Check protocol to ensure steps or ports have not been modified.

Check that valves above the tray are in the open positions.

Excessive washing. Reduce number or duration of wash steps.

Reduce concentration of detergents, such as Tween 20, if used.

Excessive signal. Too much protein loaded on the gel.

Load less protein.

Antibody concentrations too high.

Try lower concentrations of antibody.

Inadequate washing. Increase the number of washes or extend the wash times.

Non-specific binding.

Impure antibodies or other proteins reacting with similar binding sites as antigen.

Try lower concentrations of antibody or further purify antibody.

Uneven, splotchy background.

Membrane not fully or properly rehydrated.

Pre-wet membranes prior to incubation. Wet nitrocellulose membranes in water, then equilibrate with wash buffer. Wet PVDF first in methanol, then equilibrate with wash buffer.

Fingerprints, marks or damage to the membrane can cause non-specific binding artifacts.

Wear gloves and use forceps when handling membranes. Membranes are very fragile. Avoid tearing, folding or creasing membranes to minimize processing artifacts.

Areas of the blot were not fully coated with blocking agent or the membrane partially dried during incubation.

Ensure proper volumes used for each tray position. Ensure reagent lines are secured into reagent vessels and that the opaque reagent tubing reaches to the bottom of the container.

Membrane has moved into the drain ramps.

Make sure the processed membrane is at least 1.5 cm2, on the Mini Trays, or 2.5 cm2, on the Standard Trays, to deter flow into drain ramp.

High background. Inadequate blocking. Blocking agent contaminated or too dilute.

Inadequate washing between antibody steps.

Extend the time, volume or number of washes.

Choice of membrane. Avoid using nylon membranes.

Contaminated buffers. Prepare fresh buffers.

Cross-reactivity with blocking reagents.

Try a different blocking agent, such as 0.1% Tween-20, 3% BSA, 5% non-fat dry milk, casein or fish gelatin.


C a r e a n d M a i n t e n a n c e

14 Care and Maintenance

Always turn the mains power switch off and unplug the power cord before doing any maintenance.

Important Do not autoclave any component of the Processor Plus.

After each use, clean the unit with a mild detergent and water, rinse thoroughly with distilled water and allow to air dry. Never use abrasive cleansers or solvents. Do not expose the unit to solutions or vapours of aromatic or halogenated hydrocarbons, ketones, esters, alcohols (over 30%), or concentrated acids (over 25%).

Running the cleaning protocol

Run the pre-programmed cleaning protocols:

• after every protocol

• before switching to a protocol that uses different reagents

• before switching from gel staining protocols to blot processing protocols, or vice versa

• before changing the peristaltic pump tubing

• whenever the instrument has not been in use for several days

The cleaning protocol requires approximately 10 minutes to completely rinse the reagent lines.

See page 16 for directions on using the blot processing Clean protocol. See page 32 for directions on using the gel staining Clean protocol.

Staining trays

Note Residual silver stain can accumulate on the staining trays, giving them a black appearance. This black residue is inert. It does not interfere with routine staining or cause black spots on gels.

Important To prevent the accumulation of residual silver stain, thoroughly clean the staining tray after each use.

1. First wipe residual staining solution from the tray with a lint-free tissue.

2. Detach the drain line from the drain port and lift the staining tray out of the

tray support.

3. Clean the staining tray with detergent and a non-abrasive cloth. Rinse

thoroughly with distilled water.

Avoid scratching the PTFE coating on the tray. Do not use metallic instruments on the tray surface.


Care and Maintenance

Chemical Resistance

The parts of the instrument that come into contact with liquid reagents are resistant to chemicals typically used in Coomassie and silver staining and Western blot processing. Before introducing any other chemicals into the system, first test the exposed parts. Never use ketones, hot or strong acids, or hydrocarbon solvents. Refer to Table 14-1 for more information on the chemical susceptibility of components.

Table 14-1 Chemical susceptibility of Processor Plus components

Cleaning the splash guard

If gel fragments or particulate matter clogs the splash guard or drain, first disconnect the drain line from the bottom of the staining tray, then lift the tray out of the tray support. Rinse the drain tube from the bottom and run water through the splash guard from the top to remove any lodged material.

Replacing the tray gasket on the staining tray

The gasket around the staining tray supports the glass lid and creates a seal to contain reagents inside the tray (Fig 14-1). If the gasket is damaged, pull it away from the tray edge. Fit a new gasket around the tray edge, pressing the groove in the gasket over the tray edge completely around the tray.

Fig 14-1 Placement of gasket on staining tray

Instrument Component Material Susceptible to

10-port valve, exterior PVDF ketones, esters and hot acids

10-port valve gasket, internal

Fluoro rubber hot strong acids, esters, ketones and bleach

Tubing10-port valve to trayReagent lines

Pump head tubing

TygonPVC

C-flex

esters, ketones and hydrocarbonshot acids, ketones and

hydrocarbonshydrocarbons, ethers, ketones,

phenol, oils

Processing tray Styrene hydrocarbons, strong acids and bases, absolute methanol

Staining tray PTFE-coated stainless steel

long exposure to highly concentrated acids and bases

Gasket



Pump Care and Maintenance

Calibrating delivery volume

For Blot processing, see page 11. For Gel staining, see page 28.

Changing reagent lines

The valve at the back of the instrument houses ten valve ports. Ten reagent line assemblies connect to these valve ports. Replace reagent lines as they become soiled or worn.

.

Fig 14-2 10-port valve and reagent line assemblies

Each of the ten reagent line assemblies consists of two types of tubing that fit together.

Important Any modifications to the reagent line assembly will affect pump performance and calibration. Use only reagent lines supplied by Amersham Biosciences(code no. 80-6342-58). Do not detach the opaque, rigid tube from the end of the reagent line.

1. When removing the flexible end of the reagent line from the valve port, tug

the line gently just where it slips over the valve port and pull it straight out.

To avoid breaking valve ports on the 10-port valve, do not work the line

from side to side.

2. To install the new reagent lines, press the flexible end of the new line onto

the valve port.

Ten reagent line assemblies

10-port valve



Changing the peristaltic pump tubing

Change the peristaltic pump tubing when the white tubing inside the pump leaks or becomes worn.

The peristaltic pump on the rear of the instrument consists of two pump heads. The two pump heads are held together and in place by four wingnuts on the ends of four threaded studs.

Two pieces of tubing wrap around the pump heads. The four ends of the tubing that come out of the pump are shown in Fig 14-3.

Fig 14-3 The four pieces of tubing coming out of the peristaltic pump

Your selection of either the blot processing trays or the gel staining trays determines how these four pieces of tubing are connected. Table 14-2 details the connections for each tray configuration.

Table 14-2 Pump tubing connections

Keep a supply of paper towels handy to absorb any water that may run out of the tubing during disassembly.

1. To minimize contact with reagents, run the cleaning protocol to rinse the

reagent lines with distilled water before replacing the pump head tubing.

See page 16 for a description of the Clean protocol for blot processing; see page 32 for a description of the Clean protocol for gel staining.

Tubing # Blot Process Gel Staining

1 To 10-port valve To 10-port valve

2 To lid input manifold To staining tray drain

3 To waste bottle Not in use

4 To tray drain tube Not in use

1

3 2

4



2. Disconnect the pump tubing from either the blot processing tray lid (# 2 and

#4 in Fig 14-3) or the gel staining tray (#2).

3. Disconnect the short translucent tubing (#1 in Fig 14-3) from the 10-port

valve.

4. Unscrew the four wingnuts that hold the two heads of the peristaltic pump

against the rear panel.

5. Pull the outer pump head off the four threaded rods.

6. Pull the top half of the outer pump head off. Peel the pump head tubing

away from the pump shaft and discard it. This is the tubing that ends with

# 3 and #4 in Fig 14-3.

7. Place the new pump head tubing around the pump shaft (Fig 14-4).

As shown in Fig 14-4, hold the pump head with the tang end facing up. The tubing end with the QuickFit connector (#4) should protrude only 8 cm (3 in.) from the body of the pump.

Important Be sure to use the replacement tubing with the appropriate diameter.

Fig 14-4 Replacement tubing wrapped around the shaft of the outer pump

8. Press the pump head shell over the tubing, making sure not to pinch the

tubing at any point.

9. Turn the pump head over, so that the tang points away from you. Press a

piece of dampener tape over the centre slot on the rear of the pump head. See

Figure 14-5 on page 52.

Dampener tape is clear polyester film with adhesive backing. A strip with 10 rectangular pieces of dampener tape, 13 × 14 mm each, are included in replacement tubing sets. As the name implies, dampener tape muffles the sound of the pump in operation.

3

4

Tang facing up



Fig 14-5 Press dampener tape over centre slot on rear of pump head

Set this outer pump head aside while you replace the tubing on the inner pump head.

10. Pull the inner pump head off the four threaded rods. Pull the top half of the

inner pump head off, remove and discard the pump head tubing.

11. Wrap the replacement tubing around the shaft of the inner pump.

A tie wrap attaches a short segment of translucent tubing, 0.32 cm I.D.(1/8 in), to one end of the tubing assembly for the inner pump (#1 in Fig 14-6). Keep this tie wrap close to the inner pump head housing.

Fig 14-6 The assembled inner pump, with tubing in place, tang facing up

12. Press the pump head shell over the tubing, making sure not to pinch the

tubing at any point (Fig 14-6).

13. Turn the pump head over, so that the tang points away from you. Press a

piece of dampener tape over the centre slot on the rear of the inner pump

head, as shown in Figure 14-5 on page 52.

14. Align the holes in the inner pump head housing with the four threaded rods

that protrude from the rear of the unit. Slip the inner pump head over these

rods.

Placement of dampener tape

1

Tang

2



Attach the short piece of translucent tubing (#1 in Fig 14-7) to the port on the outside of the 10-port valve. Do not allow kinks to form in this tubing.

Fig 14-7 The inner pump head in place on the rear of the unit

15. Use your fingers to turn the tang on the end of the pump head shaft (Fig 14-

7) until it aligns with the tang on the end of the motor drive shaft. When the

two tangs are in alignment, the inner pump head sits flush against the unit,

with no gaps between the pump housing and the unit.

Note The pump head tang does not move easily. You will encounter some resistance when turning the tang.

16. Now slip the outer pump head on, over the four threaded rods. Turn the tang

on the end of the outer pump head until it aligns with the tang on the inner

pump head. The tang does not move easily.

17. When the outer pump head is in place, replace the four wingnuts on the ends

of the threaded rods and tighten them hand tight.

18. Connect the pump tubing to the tray/lid assembly. See Table 14-2 on page 50

for a description of the connections.

Important Always re-calibrate the pump after changing the pump head tubing.

• For blot processing, see page 11 for a description of the pump calibration.

• For gel staining, see page 28 for a description of the pump calibration.

1

2

Tang


Pre-programmed Blot Processing Protocols

Appendix A Pre-programmed Blot Processing ProtocolsIn the pre-programmed blot processing protocols, port numbers correspond to the following reagents:

Table A-1 Summary of Pre-programmed Blot Processing Protocols

Protocol 1. ECL Plus, for use with the ECL Plus Western Blotting Detection

system.

Table A-2 Protocol 1, ECL Plus

Port Reagent Description

5 Block A buffer, containing protein or detergent, that blocks unbound sites on membranes and prevents antibodies from sticking to unbound regions. Non-fat dry milk, casein, Tween 20 or BSA are often used.

6 Wash buffer Tris-buffered saline containing Tween 20 (TBST) or Phosphate-buffered saline containing Tween 20 (PBST)

1 Primary (1°) ab

2 Secondary (2°) ab

4 Enhancement reagent Avidin or stepavidin conjugate or complex. Enhances signal by adding more binding sites for detection components.

0 Air vent

Protocol Name

Application Steps Approx. Time (min.)

1. ECL Plus ECL Plus Detection Kit 24 340

2. ECL ECL Detection Kit 18 350

3. Standard Immunodetection with 1° & 2° ab 9 210

4. Enhanced Immunodetection with 1° & 2° ab and avidin products

13 285

5. Clean Clean reagent lines 9 10

Step Reagent In-Port Multiplier Time (min.)

1 Block 5 2 60

2 Wash 6 3 2

3 Wash 6 3 2

4 1° Antibody 1 1 60

5 Wash 6 2 1

6 Wash 6 2 1

7 Wash 6 3 15

8 Wash 6 3 5

9 Wash 6 3 5

10 Wash 6 3 5


P r e - p r o g r a m m e d B l o t P r o c e s s i n g P r o t o c o l s

Protocol 2. ECL, for use with the ECL Western Blotting Detection system. If

biotinylated 2° antibody is used in Step 9, then strepavidin HRP is used in Step 15.

Table A-3 Protocol 2, ECL

11 Biotinylated 2° antibody 2 1 60

12 Wash 6 2 1

13 Wash 6 2 1

14 Wash 6 3 15

15 Wash 6 3 5

16 Wash 6 3 5

17 Wash 6 3 5

18 Strepavidin HRP 4 1 60

19 Wash 6 2 1

20 Wash 6 2 1

21 Wash 6 3 15

22 Wash 6 3 5

23 Wash 6 3 5

24 Wash 6 3 5


1 Block 5 2 60

2 Wash 6 3 15

3 Wash 6 3 5

4 Wash 6 3 5

5 1° Antibody 1 1 60

6 Wash 6 3 15

7 Wash 6 3 5

8 Wash 6 3 5

9 Biotinylated 2° ab or 2° ab HRP conjugate

2 1 60

10 Wash 6 3 15

11 Wash 6 3 5

12 Wash 6 3 5

13 Enhancement (optional) 4 1 60

14 Wash 6 3 15

15 Wash 6 3 5

16 Wash 6 3 5

17 Wash 6 3 5

18 Wash 6 3 5



Pre-programmed Blot Processing Protocols

Protocol 3. Standard, for Immunodetection with primary and secondary

antibodies

Table A-4 Protocol 3, Standard

Protocol 4. Enhanced, for Immunodetection with primary and secondary

antibodies and avidin products

Table A-5 Protocol 4, Enhanced


1 Block 5 2 60

2 1° antibody 1 1 60

3 Wash 6 3 5

4 Wash 6 3 5

5 Wash 6 3 5

6 2° antibody conjugate 2 1 60

7 Wash 6 3 5

8 Wash 6 3 5

9 Wash 6 3 5


1 Block 5 2 60

2 1° antibody 1 1 60

3 Wash 6 3 5

4 Wash 6 3 5

5 Wash 6 3 5

6 Biotinylated 2° antibody 2 1 60

7 Wash 6 3 5

8 Wash 6 3 5

9 Wash 6 3 5

10 A conjugate of:Strep HRP or APAvidin HRP or AP

4 1 60

11 Wash 6 3 5

12 Wash 6 3 5

13 Wash 6 3 5


P r e - p r o g r a m m e d B l o t P r o c e s s i n g P r o t o c o l s

Protocol 5. Cleaning reagent line assemblies

Table A-6 Protocol 5, Clean


1 Distilled water or TBST 1 3 1










Pre-programmed Staining Protocols

Appendix B Pre-programmed Staining Protocols

Table B-1 Summary of Pre-programmed Gel Staining Protocols

Protocol #1. DNA silver staining, for staining 1-mm thick, unbacked DNA gels

or 0.5-mm thick, DNA gels on plastic backing (ExcelGel or CleanGel). For use with PlusOne DNA Silver Staining Kit.

Table B-2 Protocol 1, DNA silver staining

Protocol Name

Application Steps Approx. Time (min.)

1. DNA Silver DNA Gels1.0-mm unbacked gels0.5-mm backed gels

5 100

2. Protein Silver SDS and Native0.75- and 1.0-mm unbacked gels 0.5-mm backed gels Immobiline DryPlate gels

14 160

3. Protein Silver SDS and Native1.5-mm unbacked gels

14 180

4. Protein Silver IEF0.5- and 1-mm backed gels

18 300

5. Protein Coomassie SDS and Native1.0-mm unbacked gels0.5-mm backed gels

8 320

6. Protein Coomassie SDS and Native0.75-mm unbacked gels

8 250

7. Protein Coomassie SDS and Native1.5-mm unbacked gels

8 460

8. Protein Coomassie IEF0.5- and 1-mm backed gels

9 350

Step # Solution IN-port OUT-port Time (min.)

1 Fixing solution 1 8 30

2 Silver solution 3 9 30

3 Water 0 7 1

4 Developing solution 4 8 6

5 Stopping/preserving solution 5 7 30 (+ hold)


P r e - p r o g r a m m e d S t a i n i n g P r o t o c o l s

Protocol #2. Protein silver staining, for unbacked 0.75 and 1.0 mm thick SDS

and native polyacrylamide gels, or 0.5 mm thick SDS and native polyacrylamide gels on plastic backing (ExcelGel or CleanGel). Can also be used to stain Immobiline DryPlate gels. For use with PlusOne Silver Staining Kit, Protein.

Table B-3 Protocol 2, Protein silver staining

Protocol #3. Protein silver staining, for unbacked 1.5 mm thick SDS and native

polyacrylamide gels. For use with PlusOne Silver Staining Kit, Protein.




2 Sensitizing solution 2 8 30

3 Water 0 7 5

4 Water 0 7 5

5 Water 0 7 5


7 Water 0 7 5

8 Water 0 7 5


10 Stop solution 5 7 10

11 Water 0 7 5

12 Water 0 7 5

13 Water 0 7 5

14 Preserving solution 6 7 30 (+ hold)




3 Water 0 7 10

4 Water 0 7 10

5 Water 0 7 10


7 Water 0 7 1

8 Water 0 7 1



11 Water 0 7 5

12 Water 0 7 5

13 Water 0 7 5




Protocol #4. Protein silver staining: A silver staining protocol for carrier

ampholyte-containing IEF gels on plastic backing. For use with PlusOne Silver Staining Kit, Protein.


Protocol #5. Protein Coomassie staining, for 1.0 mm thick unbacked gels, or

0.5 mm thick SDS-polyacrylamide gels on plastic backing (ExcelGel or CleanGel). For use with Coomassie tablets, PhastGel Blue.

Table B-6 Protocol 5, Protein Coomassie staining


1 Fixing solution #1 7 9 45


3 5% Methanol 8 9 15





8 Water 0 9 5

9 Water 0 9 5


11 Water 0 9 1

12 Water 0 9 1



15 Water 0 9 5

16 Water 0 9 5

17 Water 0 9 5




2 Destain solution 2 9 2

3 0.02% Coomassie Blue 3 3 60







P r e - p r o g r a m m e d S t a i n i n g P r o t o c o l s

Protocol #6. Protein Coomassie staining, for 0.75 mm thick unbacked SDS-

polyacrylamide gels. For use with Coomassie tablets, PhastGel Blue.


Protocol #7. Protein Coomassie staining, for 1.5 mm thick unbacked SDS-

polyacrylamide gels. For use with Coomassie tablets, PhastGel Blue.




2 Destain solution 2 9 1.5


















Protocol #8. Protein Coomassie staining, for 0.5- or 1.0-mm thick carrier

ampholyte-containing IEF gels on plastic backing (Ampholine PAGplate or CleanGel IEF). For use with Coomassie tablets, PhastGel Blue.


Protocol #9. Clean

Table B-10 Protocol 9, Clean





4 0.02% Coomassie Blue, 0.1% CuSO4 3 3 60







1 Distilled water 0 1 0.5










B l o t P r o c e s s P r o t o c o l W o r k s h e e t

Appendix C Blot Process Protocol WorksheetProtocol #________ Processor tray Mini Standard Date__________

Membrane type________________________________ Number of Membranes______

Experiment_________________________________________________________________________________________________________________________________

Step # Solution IN-port Multiplier Time (min.)

1

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30


Gel Stain Protocol Worksheet

Appendix D Gel Stain Protocol Worksheet

Protocol #___ Date______

SSSSttttaaaaiiiinnnn Silver Coomassie

GGGGeeeellll DNA Protein Number of Gels___ Thickness______

TTTTrrrraaaayyyy 19 × 29 cm 29 × 35 cm Volume______ml


1

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30


S p e c i f i c a t i o n s

Appendix E Specifications

*This declaration of conformity is only valid for the instrument when it is:

• used in laboratory locations, and

• used as delivered from Amersham Biosciences , except for alterations described in the User Manual.

Blot Processing TraysMiniStandard

Styrene10 – 50 ml per chamber 25 –125 ml per chamber

Gel Staining Trays

19 × 29 cm 29 × 35 cm

PTFE (polytetrafluoroethylene)-coated stainless steel

125 – 200 ml 250 – 400 ml

Reagent ports 10

Programmable parametersBlot processing

Gel Staining

Input port, Volume multiplier, Time, Rock, Hold, BeepInput port, Output port, Time, Rock, Hold, Beep

Protocols 10 for Blot Processing14 for Gel Staining

Protocol key Removable, holds 1 protocol

Keypad 10-key membrane switch

Display Liquid crystal display (LCD), two lines, 16 characters per line

Alarm Programmable, audible beep

Operating temperature 4–40 °C

Relative humidity Less than 80% for 4–31 °C, decreasing linearly to 50% for 31–40 °C

Line voltage 115 V or 230 V

Frequency 60 Hz or 50 Hz

Maximum power 60 W

Dimensions (h × w × d) 22.5 × 40 × 51 cm (with Mini or Standard tray)21 × 30 × 47 cm (with 19 × 29 cm tray)21 × 35 × 47 cm (with 29 × 35 cm tray)

Weight, maximum 12.0 kg (with 29 × 35 cm tray)

Safety certifications *CE, UL, CSA


Customer Service Information


Technical Service and Repair

Amersham Biosciences offers complete technical support for all our products. If you have any questions about how to use this product, or would like to arrange to repair it, please call or fax your local Amersham Biosciencesrepresentative.

Important repacking instructions1. Request a copy of the Amersham Biosciences “Health and Safety

Declaration” Form before returning the item. No items can be accepted for

servicing or return unless this form is properly completed.

2. The rocker finger must be in the down position before repacking the unit,

but it must never be moved manually. To reposition it, turn the power off and

then on. During the diagnostic cycle, the rocker finger moves along its range

of motion. Turn the power off when the finger is in the down position, then

pack the instrument.


C u s t o m e r S e r v i c e I n f o r m a t i o n

Ordering Information

Qty Code No.

Processor Plus, Blot Processing System

Hoefer Processor Plus, base unitIncludes pump and reagent lines, protocol key

1 80-6444-04

Blot Processing Tray PackIncludes tray support, disposable Mini and Standard trays, lid with tubing, four reagent bottles and a rack, waste bottle cap (38-430), spirit level, lid stand, Membrane Processing Technical Manual

1 80-6444-23

Replacement Parts for Blot Processing

Blot Processing tray, disposable Mini (3/pkg) 1 pkg 80-6444-42

Blot Processing tray, disposable Standard (3/pkg) 1 pkg 80-6444-61

Blot Processing Tubing Set, completeIncludes lid manifold with valves, pump tubing, dampeners, couplings, and reagent lines

1 kit 80-6448-22

Reagent line Set 10 80-6342-58

Pump tubing Set 1 set 80-6448-03

Reagent bottles, 4/pkg 1 pkg 80-6448-79

ECL and ECL Plus Reagents

ECL Western blotting reagentsFor 1000 cm2 membraneFor 2000 cm2 membraneFor 4000 cm2 membrane

1 kit1 kit1 kit

RPN2109RPN2209RPN2106

ECL Western blotting analysis system 1 RPN2108

ECL Plus Western blotting detection reagents For 1000 cm2 membrane 1 RPN2132

Protein Membranes

Hybond-C pure20 × 20 cm, sheets20 cm × 3 m, roll30 cm × 3 m, roll

10 11

RPN2020WRPN203WRPN303W

Hybond-P 20 × 20 cm, sheets30 cm × 3 m, roll

10 1

RPN2020FRPN303F

Hybond ECL6 × 8 cm, sheets20 × 20 cm, sheets30 cm × 3 m, roll

50101

RPN68DRPN2020DRPN303D

Hybond-C extra20 × 20 cm, sheets20 × 50 cm, sheets20 cm × 3 m, roll30 cm × 3 m, roll

10511

RPN2020ERPN3050ERPN203ERPN303E



Processor Plus, Gel Staining System

Hoefer Processor Plus, base unitIncludes pump and reagent lines, protocol key

1 80-6444-04

Gel Staining Tray Pack (19 x 29 cm tray)Includes tray support, PTFE-coated staining tray, lid, magnets, gasket, level and Gel Staining Protocol Guide

1 80-6444-80

Gel Staining Tray Pack (29 x 35 cm tray)Includes tray support, PTFE-coated staining tray, lid, magnets, gasket, level and Gel Staining Protocol Guide

1 80-6445-18

Replacement Parts for Gel Staining

Staining Tray, PTFE-coated, 19 x 29 cm 1 80-6444-99

Glass lid, 19 x 29 cm 1 80-6341-82

Tray gasket, for 19 x 29 cm tray 1 80-6341-44

Tray support, for 19 x 29 cm tray 1 80-6344-29

Staining Tray, PTFE-coated, 29 x 35 cm 1 80-6445-37

Glass Lid, 29 x 35 cm 1 80-6341-63

Tray gasket, for 29 x 35 cm tray 1 80-6341-25

Tray support, for 29 x 35 cm tray 1 80-6345-05

Splash Guard for delivery port 1 80-6342-01

Magnets 4 80-6342-20

Gel Staining Tubing Set, completeIncludes tubing for tray and pump, QuickFit couplings, reagent lines and dampeners

1 set 80-6445-56

Reagent line Set 10 80-6342-58

Pump Tubing Set 1 set 80-6448-03

Staining Kits

PlusOne DNA Silver Staining Kit 1 17-6000-30

PlusOne Silver Staining Kit, Protein 1 17-1150-01

Coomassie tablets, PhastGel Blue 40 17-0518-01

Additional Replacement Parts and Accessories

Protocol key 1 80-6342-77

Power cord, 115V 1 80-6106-03

Power cord, 230V 1 80-6230-10

Serial printer cable + adaptor 1 80-6427-70

Spirit Level 1 80-6087-60

Protocol Guide: Automated Silver and Coomassie Staining of Polyacrylamide Gels

1 80-6343-34

Technical Manual: Membrane Processing 1 80-6447-27

Quick Reference Card 1 80-6447-08

Processor Plus User Manual 1 80-6446-89

Qty Code No.


iiiinnnnddddeeeexxxx

NNumerics10-port valve 2

Aadding a step 19, 20, 36, 37arrow keys 5autoclave, avoiding 47

BBeep 15, 18, 31, 34blot processing 8–25

clean protocol 16leveling tray 9, 23manual pump operation 22preparation 13print protocol 15protocols 54–57pump calibration 11reagent volume 18, 23setup parameters 22–25step variables 17tray sizes 3, 23worksheet 63

Cclean protocol

blot processing 16gel staining 32

cleaning exterior and trays 47

Ddeleting a step 20, 21, 36, 37

EEDIT/SET key 5, 7editing

a protocol in progress 20, 36protocol name 17, 33step variables 17, 33

end-of-step options 17, 33

Ggel staining 26–40

clean protocol 32leveling tray 27, 39manual pump operation 38preparation 30print protocol 31, 40protocols 58–62pump calibration 28reagent volume 34, 39setup parameters 38–40step variables 33tray sizes 3, 26, 39worksheet 64

HHold 18, 34

Kkeypad 2, 5

LLCD 2level tray position 15, 32

Mmagnets 3Main menu 5manual pump operation


multiplier, for minimum volume 18, 34

Ooperating software navigation 5–7

PPAUSE mode 15, 32peristaltic pump 2

tubing 2, 50pivot ball assembly 2Port In 18, 34Port Out 34power outage 21, 37, 43preparing

gels 30membranes 13

printing 1, 4protocol


Processor Pluscomponents 2specifications 65

Protocol 5Clean, blot processing 16

Protocol 9Clean, gel staining 32

Protocol key 1, 2slot 2use 41


protocolsadding a step 19blot processing 54–57cleaning 16, 32deleting a step 20displaying volume 23editing 19gel staining 58–62interrupting 15, 31printing


printing volume 24start operation 14, 15, 30, 31termination 15, 32

pump calibrationblot processing 11gel staining 28

pump head tubingchanging 50

PVDFchemical susceptibility 48

QQuick-Fit couplings 9, 27

Rreagent lines 2reagent tubing assembly 49

changing 49cleaning 16, 32

Recover, tray position 15, 32Rock 18, 34rocker finger 2

Sself-diagnostic cycle 5Setup 1


Setup 2blot processing 23gel staining 39





Setup 7blot processing 11

Setup menus 7

Setup parametersblot processing 22–25gel staining 38–40

silver stain residue 47software navigation 5–7specifications 65spirit level 2splash guard 3

cleaning 48step variables

blot processing 17gel staining 33Table


stopping a protocol 15, 32

Ttray gasket replacement 48tray level


tray sizesand volume 23, 26, 39blot processing 23gel staining 26, 39

troubleshooting 43–45

Vvalves, on blot processing lid 13volume delivery

blot processing 23gel staining 30, 39

Wworksheet



Amersham Biosciences UK Limited Amersham Place Little Chalfont Bucks HP7 9NA England Amersham Biosciences AB SE-751 84 Uppsala SwedenAmersham Biosciences Inc 800 Centennial Avenue PO Box 1327 Piscataway NJ 08855 USAAmersham Biosciences Europe GmbH Postfach 5480 9 D-79021 Freiburg

ECL, ExcelGel, CleanGel, Hoefer, Hybond, PlusOne, and Rainbow are trademarks of Amersham Biosciences Limited or its subsidiaries.Amersham and Amersham Biosciences is a trademark of Amersham plc

Coomassie is a trademark of ICI plcNalgene is a trademark of Nalge Nunc InternationalTween is a trademark of ICI Americas, Inc.

All goods and services are sold subject to the terms and conditions of sale of the company within the Amersham Biosciences group which supplies them. A copy of these terms and conditions is available on request.

© Amersham Biosciences 1999—All rights reserved.

Printed in USA

HB, Gel Staining

Documents

protocol key

blot processing tray

protocol15cleaning protocol

protocol31cleaning protocol

gel staining tray

print volume of protocol

display protocol volume

tray level