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RMMS FOR MICROBIAL IDENTIFICATION
Company
Product
Name
Scientific
Method
App lic ations
Time to
ResultThroughput
Sample Size
or TypeSensitivity
Organism
LibrariesWorkflow
Abbott
Laboratories,
Ibis Biosciences
Division
PLEX-ID System
Note that this
RMM is currently
being redesigned
and is currently
unavailable.
Genotyping by Multi
Loci Base
Composition
(MLBC) analysis
using PCR and
mass spectrometry
Identification
6-8 hr250 every 24
hr
40-160 uL of
nucleic acid
sample
10 copies per
gene
Bacteria, fungi and
viruses. > 750,000
microbial signatures
with detection down to
the strain level
DNA is extracted from cells originating
from colonies or from liquid samples.
The DNA sample is applied to specific
wells of a 96 well-plate, preloaded
with sequence primer pairs. After PCR
amplification, the plate is loaded on
the PLEX-ID system where pico-liter
amounts of the PCR products are
analyzed in an electrospray TOF
mass spectrometer. The base
composition for all amplicons present
are determined and searched against
a database to determine the
organisms present. This multilocus
approach provides the ability to
distinguish between closely related
species as well as single nucleotide
polymorphisms or mutations. There isno requirement for additional staining,
culturing or enrichment steps. Simple
mixtures of organisms can also be
analyzed. Not for use in diagnostic
procedures.
Applied
Biosystems
MicroSEQ
PCR and gene
sequencing
Identification
4-5 hr
80 per day.
Higher with
greater
capacity
capillaryanalyzers.
Cells from
colonyNot applicable
>1800 bacteria
>90 Mycoplasma
>1100 yeast and mold
DNA is extracted from cells originating
from isolated colonies. Amplify target
16S rDNA for bacteria or the D2
region of large-subunit rDNA for fungi
using PCR. Perform sequencing of the
PCR amplicons, resulting in DNA
fragments of various sizes ending in
different nucleotides/dyes. A genetic
analyzer separates the fragments by
size and a laser detects thefluorescence color from each dye,
producing a full gene sequence of the
target DNA. The resulting sequence is
compared with an internal database of
known sequences. No Gram staining
is required.
Battelle
REBS
Raman
spectroscopy
Identification and
enumeration
3 min160 every 8
hr
Cells from
colony, liquid
medium,
product/raw
material,
surfaces
1 cell is
identified and
quantified
Alcaligenes,
Pseudomonas,
Brevundimonas,
Candida, E. coli,
Bacillus, Ralstonia,
vegetative and spore
forms.
Sample material is retained on a
supported film. The area is examined
for microscopic particulates using
Raman spectroscopy and a spectral
signature is provided for each
particulate. The spectral signatures
are statistically correlated to a library
of known microorganisms. No need
for Gram staining. Mixed cultures and
be identified and enumerated.
Non-destructive for further analysis.
Biolog
OmniLog;
MicroStation;
MicroLog
Growth-based;
carbohydrate
utilization.
Identification
2-72 hr 50 per dayCells from
colonyNot applicable
>1226 bacteria and
>885 yeast/mold (GEN
II cards)
>1000 bacteria (GEN III
card)
Cells from isolated colonies are used
to prepare a microbial suspension,
which is then added to specific test
cards containing a variety of
carbohydrates and a colorless
tetrazolium violet dye. If growth
occurs, the dye turns violet in color.
The resulting color patterns are
compared with an internal library.
Gram staining is required when using
GEN II test cards for bacteria, yeast
and mold. No Gram stain is required
for the GEN III bacterial card (ID's
both Gram + and - bacteria).
BD Diagnostic
Systems
Phoenix
Growth-based;
biochemical
utilization and
antibiotic
3 hr 100 per dayCells from
colonyNot applicable >225 bacteria
Cells from isolated colonies are used
to prepare a microbial suspension,
which is then added to specific test
cards containing substrates (in wells)
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susceptibility
Identification
for biochemical utilization. Color or
fluorescence changes in each well are
compared with an internal library.
Gram staining is required to determine
the correct test card to use.
bioMrieux
Vitek 2 Compact
Growth-based;
biochemical and
carbohydrate
utilization.
Identification
2-18 hr 30-60 per dayCells from
colonyNot applicable
>285 bacteria
>48 yeast
Cells from isolated colonies are used
to prepare a microbial suspension,
which is then added to specific test
cards containing substrates for
enzymatic utilization, carbohydrate
acidification and other tests. Color or
turbidity changes in each well are
measured every 15 minutes and
results are compared with an internal
library. Gram staining is required to
determine the correct test card to use.
bioMrieux
DiversiLab
PCR
Identification
4 hr
13 samples
per chip in 4
hr. Each
additional
chip (13
samples)
every 1 hr.
Cells from
colonyNot applicable
>168 bacteria
>1100 strain or
sub-species patterns
DNA is extracted from cells originating
from isolated colonies. Amplify target
DNA (noncoding repetitive DNA
sequences) using PCR. PCR products
are then separated using
electrophoresis in a microfluidics chip
and the resulting patterns are
compared to an internal library.
bioMrieux
Vitek MS
MALDI TOF mass
spectrometry
Identification
2 min480 every 8
hr
Cells from
colonyNot applicable
508 bacteria
78 fungi
Cells from isolated colonies or liquid
medium are added to a stainless steel
target plate and allowed to dry. A
UV-absorbing matrix is added and the
cells are ionized by a laser. The
ionized particles are accelerated in an
electric field and enter the time of
flight (TOF) tube, where protein and
peptide molecules are separated
according to their mass to charge
ratio. The resulting MALDI-TOF mass
spectrum is compared with an internal
database. Gram staining is not
required. Mixed culture ID possible.
Bruker Daltonics
MALDI Biotyper
MALDI-TOF mass
spectrometry
Identification
1-2 min 30-60 per hr
Cells from
colony or liquid
sample
Not applicable
> 2,000 species and >
4,010 strains of
bacteria, yeast and
mold
Cells from isolated colonies or liquid
medium are added to a stainless steel
target plate and allowed to dry. A
UV-absorbing matrix is added and the
cells are ionized by a laser. The
ionized particles are accelerated in an
electric field and enter the time of
flight (TOF) tube, where protein and
peptide molecules are separated
according to their mass to charge
ratio. The resulting MALDI-TOF mass
spectrum is compared with an internal
database. Gram staining is not
required. Mixed culture ID possible.
Bruker Daltonics
TENSOR
27+HTS-XT
Fourier TransformInfrared (FT-IR)
Spectrometry
Identification
Minutes
Continuous
sampling on
96 and 384
plate formats
Cells from
colonyNot applicable
Yeast, bacilli,
Coryneforms,
Micrococci,
Staphylococci, Bifido,
Clostridia,
Pseudomonads,
Lactobacilli,
Acetobactereaceae
Microorganisms are harvested from
the cultivation medium, suspended in
water and then transferred on a
special IR-transparent, reusable
sample plate. The measurement is
performed after drying of the samples.
The dried biofilm is then analyzed by
the FT-IR microplate reader. The
FT-IR spectrum is then compared with
an internal library.
ceeram
ceeramTools
Genotyping Kits
LP
MLVA (Multi Locus
VNTR Analysis)
Genotyping
2 days100 samples
in 2 days
DNA or culture
cell
Not applicable
Legionella
pneumophila,
Staphylococcus aureus,
Pseudomonas
aeruginosa
MLVA is a PCR based typing method
that relies on the inherent variability
found in many regions of repetitive
DNA called VNTR (Variable Number
Tandem Repeat) which represent
sources of polymorphisms. The MLVA
assay for L. pneumophila examines
12 loci and the assay for S. aureus
and P. aeruginosa examines 16 loci.
Thus, each isolate is defined by a 12
or 16-digit numeric code
corresponding to the number of
repeats at each VNTR. Efficient
amplification of the markers is
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performed in a single or 2 multiplex
PCR reactions. An ABI sequencer is
required to analyze the amplicons.
Dupont Qualicon
Riboprinter
Ribotyping of DNA
fragments
Identification
8 hr 32 per dayCells from
colonyNot applicable
>1,440 bacteria
>8,500 strain or
sub-species patterns
DNA is extracted from cells originating
from isolated colonies. The DNA is cut
into fragments using a restriction
enzyme, which are then separated
according to size. The DNA is
immobilized on a nylon membrane,
denatured to produce single-stranded
DNA, and then hybridized with a DNA
probe (derived from an E. colirRNA
operon). An antibody-enzyme
conjugate is bound to the probe and a
chemiluminescent agent is added,
resulting in a banding pattern that is
compared with an internal database.
Fully automated; no Gram staining
required.
Greiner Bio-One
CytoInspect
PCR and microarray
analysis
Mycoplasma
detection and
identification
5 hr 100 per day
Detection of > 90
species of Mycoplasmaand identification of 40
Mycoplasma species
A microarray based test kit for the
detection and identification of
mycoplasma species in cell cultures
and other biological materials. DNA is
extracted and PCR performed using
primers specific for conserved and
species-specific regions of the16S-23S rRNA intergenic transcribed
spacer (ITS) of Mycoplasma DNA.
The fluorescently labeled fragments
are then hybridized to the microarray
chip. The chip contains probes for
both species-specific targets and a
universal probe for all Mycoplasma.
MIDI
Sherlock MIS
Detection of fatty
acids.
Identification
Standard
method (2 hr);
Instant FAME
method (1,200 bacteria
>200 yeast/mold
Fatty acids are extracted from cells
originating from isolated colonies. The
fatty acids are purified and analyzed
using gas chromatography. The
resulting GC chromatogram is
compared with an internal library. No
Gram staining is required.
Pathogenetix, Inc.
Genome
Sequence
Scanning
Fluorescent tagging
of single molecules
of genomic DNA
Identification,
analysis of microbial
mixtures,
epidemiology,
contamination
source tracing
3.5 - 4.5 hr50 samples
per 24 hr
107- 10
9cells
in 0.1 - 5 mL of
culture or from
colony
0.1% of
microbial target
mixed with other
bacteria
1,000+ organisms
including bacteria,
yeast, mold, and
Mycoplasma. Library
can be customized.
Rack of tubes with suspended cells is
inserted into an automated Sample
Preparation Module to be processed
in parallel. The Module isolates and
purifies DNA, specifically digests it,
tags DNA with fluorescent probes, and
elutes the samples for consecutive
measurement with the Detector
Module. The fragments of genomic
DNA are stretched and fluorescent
signatures generated by the probes
are measured one-by-one. Each
signature is compared with internal
database to identify an isolate or
elucidate the composition of microbial
mixture.
rap.ID
Bio Particle
Explorer BPE
Viable Staining and
Imaging LED
Raman
Spectroscopy
Identification and
enumeration
3-10 min
> 150
samples per
8 hr
Cells from
colony, liquid
medium,
product/raw
material,
surfaces
1 cell is
identified and
quantified
> 30 bacteria; can add
new entries
The sample material is collected on
metal foil. Viability staining and
automated image analysis using dark
field illumination detects viable particle
quantity, shape, and size for particles
ranging from 0.5 m and larger.
Raman spectroscopy is then
performed on each viable particle, and
a spectral signature is provided. The
spectral signatures are statistically
correlated to a library of known
microorganisms. Non-destructive for
further analysis.
Thermo Scientific
Nicolet
Fourier Transform
Infrared (FT-IR)
Spectrometry
Identification
2 hr 350 per dayCells from
colonyNot applicable Bacteria
Cells from isolated colonies are
incubated in liquid medium for 18 hr,
followed by washing, centrifugation
and resuspension in distilled water. 50
uL of the suspension is transferred
onto a special IR-transparent,
reusable sample plate and dried at
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40-45C under vacuum to create a
biofilm. The biofilm is then analyzed
and biomolecules such as proteins,
lipids, carbohydrates and DNA/RNA
are identified and quantified. The
FT-IR spectrum is then compared with
an internal library.
RMMS FOR QUALITATIVE ANALYSIS
Company
Product
Method
App lic ations
Time to
ResultThroughput
Sample Size
or TypeSensitivity
Organisms
DetectedWorkflow
BioLumix
BioLumix
System
Growth-based; CO2
detection and
selective growth
Detection of specific
organisms;
detection of
microbial growth
8-48 hr
32 tests at a
single
temperature
per
instrument
(32
instruments
can be used
with one
computer)
0.1-1.0 mL1 cell after
enrichment
Total aerobic count,
yeast & mold, coliforms,
E. coli, lactic acid
bacteria,
Enterobacteriaceae,
Salmonella,
Pseudomonas,
Staphylococcus
The test sample is added to unique
vials that contain a selective medium
and a dye. Changes in color or
fluorescence, expressed as light
intensity units, are detected by the
optical sensor and represent growing
microorganisms. The total aerobic
count and total yeast/mold vials detect
microbial growth by monitoring the
generation of CO2, and can be used
to screen for an estimation of
organisms in a test sample that are
above or below a certain quantitative
specification. Liquid and diluted solid
samples can be assayed.
Applied
Biosystems
MycoSEQ
PCR
Mycoplasma
detection
< 5 hr
100 l to 10 ml
of cell culture
sample
< 10 CFU or
copy
equivalent/ml
Detection of > 90
Mycoplasma species
The sample is added to a centrifuge
tube and Mycoplasma is concentrated
in a pellet. The Mycoplasma is
enzymatically lysed and purified using
magnetic beads and washing steps.
Real time PCR is performed and the
Mycoplasma target sequence is
detected via use of amplification plots
and melt curve analysis. If a positive
result fir Mycoplasma is obtained, the
purified DNA can be used in the
MicroSEQ for subsequent
identification.
Bactest
Speedy Breedy
Respirometry,
pressure sensing
Detection of
microbial growth;sterility testing;
presence of specific
organisms
4-20 hr 2-4 per day Up to 50 mL1 CFU after
enrichment
Aerobes, facultative
anaerobes, anaerobes,
and microaerophilicbacteria; yeast
The sample is transferred into a
disposable vessel containing a
general or selective medium. The
vessel is then placed into the portable
respirometer and incubated to
encourage microbial growth. Detection
of metabolic activity is determined by
pressure transients relating to
gaseous exchanges within the closed
culture vessel as a result of microbial
respiration. Continuous data collected
is analyzed in real time, and detection
algorithms process the pressure
transients to alert when significant
changes have taken place. The
instrument measures both positive
and negative pressure meaning that
monitoring can be performed on a
range of microbial processes reacting
to differing conditions within the
culture chamber. The system is
applicable for pharmaceuticals, raw
materials, food and beverage,
veterinary, household and hygiene
products.
BD Diagnostic
Systems
BACTEC
Growth-based; CO2
detection
Detection of
microbial growth;
sterility testing
8-48 hr
240-1200 per
incubation
period
1-10 mL or gm1 CFU after
enrichmentBacteria, yeast, mold
Samples are added directly to bottles
of liquid culture media and incubated
in the system. During microbial
growth, CO2in the closed container
accumulates and is detected by a
fluorometric sensor. The system
automatically monitors the sensor
every 10 minutes, and the generation
of CO2indicates the presence of
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growing microorganisms.
bioMrieux
Bactometer
Growth-based;
measurement of
electrical impedance
Detection of
microbial growth
6-48 hr
64-512
samples per
incubation
period
1 mL1 CFU after
enrichmentBacteria, yeast, mold
Samples are added to media in a
specialized holder that also contain
two electrodes at the bottom of each
sample test well. The holder is placed
in an incubator and continuously
monitored. Growth is detected by
monitoring the movement of ions
between electrodes (conductance), or
the storage of charge at the electrode
surface (capacitance).
bioMrieux
BacT/ALERT 3D
Dual-T
Growth-based; CO2
detection
Detection of
microbial growth;
sterility testing
24-96 hr
480-1440 per
incubation
period
1-10 mL or gm1 CFU after
enrichmentBacteria, yeast, mold
Samples are added directly to bottles
of liquid culture media and incubated
in the system (one of two
temperatures). During microbial
growth, CO2in the closed container
accumulates and diffuses into a
colorimetric sensor at the base of the
bottle. Hydrogen ions interact with the
sensor resulting in a decrease in pH,
causing the sensor to change to a
yellow color. The system automatically
monitors the sensor every 10 minutes,
and the generation of CO2indicates
the presence of growing
microorganisms.
BIOTECON
Hygiene
Screening
System
PCR
Bacterial detection
90 min 160 per dayCells from
colony
Corynebacterium,
Staphylococcus,
Macrococcus,
Micrococcus, Kocuria,
and Kytococcus
Bacterial colonies growing on agar
plates are suspended in buffer, and
the suspension is placed in a reaction
tube. Primers, fluoresence resonance
energy transfer (FRET) probes and
Taq polymerase are added to the
reaction tube. PCR amplification and
detection is carried out in a Roche
Diagnostics LightCycler 2.0
Carousel-Based System. The
amplification cycles are monitorined
via fluorescence, and melting curves
are used to determine what target
sequences/organisms are present.
CCM MuScan
Innosieve
Diagnostics Test
Kits
Viability staining and
solid phase
cytometry;
fluorescence
microscopy
Bioburden of raw
material andin-process samples,
finished product,
EM, water, sterility
testing
10-65 minutes100 per 8
hours
Filterable
samples; 1 uL
to 1L
1 - 105cells
Bacteria, yeast, fungal
spores
The test sample is
filtered/concentrated through a
0.45um micro sieve. Microorganisms
are retained on the membrane and
subsequently labeled with a viability
stain and/or a species-specific stain.
Available are total viable count, total
live-dead ratio, total species-specific
count and total species-specific viable
count. After staining a scanning period
using MuScan digital fluorescent
microscopy at specific excitation and
emission wavelengths is performed.
The image processing software
analyzes fluorescent objects on size,
shape and fluorescent signals of all
microbes. The assay can be
user-customized with different types of
fluorescent stains, labeled antibodies,
DNA-probes, etc., depending on the
organism(s) to be detected. Examples
of test kits for specific organisms
include Legionella, Salmonella,
Listeria, Chronobacter and E. coli. The
method is non-destructive such that
detected microbes can be
subsequently cultured.
ceeram
ceeramTools
RT-PCR Real
Time Detection
Kits
Real time RT-PCR
Viral detection
5 hrs
Depends on
thermocycler
used
2 gr digestive
tissue, 25 gr
fruit or
vegetable, 1 L
water
5 genome
copies
Norovirus GI, GII;
Hepatitis A, E;
Enterovirus;Adenovirus; Sapovirus;
Aichivirus; Rotavirus;
Circovirus; Brachyspira;
Giardia;
Cryptosporidium
Elution, concentration, extraction,
purification, amplification,
quantification and interpretation. Any
matrix (food, environmental, health)
may be assayed, including shellfish,
vegetable, herbs and spices, water,
sludge and surfaces. The detection
kits can be used with most PCR
instrumentation.
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Celsis
International Ltd.
Celsis Advance
System using
RapiScreen
ATP
bioluminescence
Bioburden of water,
raw materials,
in-process samples,
Microbial Limits
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used to run an assay contain precise
amounts of LAL reagent, chromogenic
substrate and control standard
endotoxin (CSE). Pipette 25 L of a
sample into each of the four sample
reservoirs of a cartridge. The reader
draws and mixes the sample with the
reagents. After mixing, the optical
density of the wells is measured and
analyzed against an internally-
archived standard curve.
Charles River
Laboratories
EndoSafe PTS
Gram ID
LAL assay
Rapid Gram staining
3-7 min4 every 3-7
min25 L Gram + and - bacteria
Using a loop, place an isolated colony
into 2-3 mLs of saline or LRW
(concentration should be 0.5
McFarland equivalence turbidity
standard units). Add 25 l each of 4
samples/organisms to 4 channels in
the disposable cartridge. Results for
Gram +, Gram , or yeast/mold are
displayed on the screen of a portable,
handheld spectrophotometer.
Charles River
Laboratories
EndoSafe PTS
Glucan Assay
LAL assay
Detection of glucan
30 min 2 per hr 25 L 10-1,000 pg/mLGlucans from yeast and
mold
Portable, handheld spectrophotometer
that utilizes disposable cartridges.
Rapid, in-process test designed for
investigational purposes to detect
(1,3)--D glucans from the cell wallsof most yeasts and molds.
Dupont Qualicon
BAX System Q7
PCR
Detection of
microorganisms
1.5-2.5 hr; 48
hr for
yeast/mold
96 per
1.5-2.5 hr10-50 L
Salmonella, Listeria
monocytogenes,
Camplyobacter
jejuni/coli, E. coli
O157:H7, Enterobacter
sakazakii,
Staphylococcus aureus,
yeast and mold, Vibrio
Samples are enriched in media to
provide enough DNA for analysis and
to eliminate false positives from dead
cells. The enriched samples are
heated in a lysis solution to release
DNA. PCR tablets, which contain all
the reagents necessary for PCR plus
fluorescent dye, are hydrated with
lysed sample and processed in the
cycler/detector. PCR amplifies a DNA
fragment that is specific to a target
organism. The amplified DNA
generates a fluorescent signal, and
results are displayed as positive ornegative results. The system uses
SYBR Green, Taqman and Scorpion
probes, facilitating the detection of
multiple species in a single sample.
Greiner Bio-One
CytoInspect
PCR and microarray
analysis
Mycoplasma
detection and
identification
5 hr 100 per day
Detection of > 90
species of Mycoplasma
and identification of 40
Mycoplasma species
A microarray based test kit for the
detection and identification of
mycoplasma species in cell cultures
and other biological materials. DNA is
extracted and PCR performed using
primers specific for conserved and
species-specific regions of the
16S-23S rRNA intergenic transcribed
spacer (ITS) of Mycoplasma DNA.
The fluorescently labeled fragments
are then hybridized to the microarraychip. The chip contains probes for
both species-specific targets and a
universal probe for all Mycoplasma.
Hitachi
BioMAYTECTOR
ATP
bioluminescence
Detection of
microorganisms
90 min5-6 samples
per 8 hr
Volumetric air
sample (e.g., 1
cubic meter)
1 cell (1
attomole ATP)
Bacteria, bacterial
spores
The system utilizes a highly-sensitive
ATP measuring instrument (100 to
200 times higher sensitivity than
conventional ATP methods) for
detecting airborne bacteria. A unique
spore germination induction
processing method is used to detect
spores. A 1 cubic meter of air is
collected within 10 minutes and
particles are deposited onto a
collection carrier. ATP from dead
bacteria are eliminated while ATPfrom viable bacteria are extracted. An
optical detection system measures
light intensity within 90 minutes.
Hyglos
ELISA
3.5 hrs96 samples
every 3 hrs100 l 0.05-500 EU/mL Endotoxin
Uses a microplate that is pre-coated
with a phage-derived receptor protein
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EndoLISADetection of
endotoxin
which has a high affinity and
specificity for the conserved core
region of LPS (endotoxin). When the
sample matrix is added to the
microplate, the LPS is bound to the
phage protein. Any sample matrix with
potentially interfering components is
then removed by a washing step. The
subsequent detection by recombinant
Factor C and a fluorescence substrate
is left unaffected by inhibitors,
facilitating a reliable quantification ofendotoxin in the sample.
Innosieve
Diagnostics
Real-Time Q-PCR
Detection Kits
Q-PCR
Bacterial detection
3-5 hrs (15-25
cycles)
Depends on
thermocycler
used
25 g
< 1 cell / 25 g
(after
pre-enrichment)
< 10 cells / g or
10
non-degraded
genomes (no
enrichment)
Campylobacter jejuni,
Clostridium perfringens,
Cronobacter sakazakii,
E. coli, Listeria innocua,
Legionella
pneumophila,
Legionella spp., Listeria
monocytogenes,
Salmonella sp.,
Staphylococcus aureus,
Vibrio alginolyticus,
Vibrio cholerae, V.
cholerae tox, Vibrio
parahaemolyticus,Vibrio vulnificus, MRSA
Taqman-based Q-PCR detection kits.
When necessary, use pre-enrichment
of the sample in an appropriate
(selective) medium (1:10 w/v: 25 g
sample + 225 ml medium) for 20- 24
hours at appropriate temperature (the
actual method is provided for each
test kit). This is followed by filtration of
100 ml with an appropriate membrane
system (e.g., 0.45 m, cellulose
nitrate filter). The detection kits can be
used with most PCR instrumentation.
JMAR
BioSentry
Light scattering
Detection of water
pathogens
2 minutesContinuous
monitoringProcess water 600 CFU/mL
Cryptosporidium,
Giardia, E. coli,
Salmonella, Shigella,
Pseudomonas,
Legionella, other
rod-shaped bacteria
The fully automated instrument
functions as a real-time, continuous
flow water-monitoring system. As the
slip stream passes through the flow
cell, it also passes through a laser
beam. When particles 0.4 microns to
10 microns in size are present, a
specific multi-angle light-scatter
pattern will be captured by the units
photodetector. This pattern is then
analyzed and compared to the
Bio-Optical Signatures database using
proprietary algorithms. From this
analysis, relative concentration is
calculated and detected particles are
classified as a bacteria, spore,
protozoan or unknown. The system
does not provide viability data as it
cannot differentiate between live and
dead microorganisms.
Lonza
MycoAlert
ATP
bioluminescence
Mycoplasma
detection
20 min 24 per 8 hr
100 L of
culturesupernatant
< 50 CFU/mL
> 90 species of
Mycoplasma
Viable Mycoplasma are lysed and the
organism's enzymes react with the
MycoAlert Substrate catalyzing the
conversion of ADP to ATP. By
measuring the level of ATP in a
sample both before and after the
addition of the MycoAlert Substrate
a ratio can be obtained which is
indicative of the presence or absence
of Mycoplasma. If these enzymes are
not present, the second reading
shows no increase over the first, while
reaction of mycoplasmal enzymes
with their specific substrates in the
MycoAlert Substrate, leads to
elevated ATP levels.
Micro
Identification
Technologies
Light scattering
Detection ofmicroorganisms
10 min 48 per 8 hr Cells from
colony
10-50 cells
E. coli,
Cryptosporidium,
Giardia
Cells from an isolated colony are
suspended in filtered water in a
sample vial and placed into the
instrument. 35 photo detectors in five
concentric arcs that surround the
sample vial collect Mie scattering light
intensities that are generated when a
cell intersects a red laser beam. The
shape, size and internal/external
structures of microorganisms will
provide a unique scattering signature,
which are compared with an internal
database.
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Merck Millipore
MilliPROBE
PCR
Mycoplasma
detection
4 hr 24 per day
Up to 20 mL of
filterable
sample
< 1-10 CFU/mL
10,000 copies
of Mycoplasma
rRNA
> 90 species of
Mycoplasma
Introduce sample into the
centrifugation assay device to
separate eukaryotic cells from
Mycoplasma. Lysis reagent and
primers for the target ribosomal RNA
sequence is added. Magnetic particles
capture and purify the RNA target.
The purified RNA is added to a
96-well plate, in addition to
amplification mix and enzyme reagent.
Real-Time Transcription Mediated
Amplification (TMA) is performed, andthe fluorescence signal from a
molecular torch probe increases as
amplification occurs. If the
Mycoplasma rRNA target is amplified,
a positive result is reported.
Pall Corporation
Pallchek Rapid
Microbiology
System
ATP
bioluminescence
Sterility testing,
bioburden (product
and surface),
non-sterile product
release, biological
indicator,
preservative
effectiveness,
sanitization
monitoring
1 min
following
24-48 hr
enrichment
and
membrane
filtration steps
100 per day
1-300 mL.
Protocol
adapted for
solids and
non-filterable
samples.
1 CFU after an
enrichment of
24 to 48 hr
Bacteria (aerobic and
anaerobic), yeast and
mold
Samples (filterable or non-filterable)
are enriched in growth media.
Incubated media is then filtered.
Microorganisms collected on the
membrane are lysed with extractant.
Luciferin and luciferase enzyme is
added and light is detected with the
Pallchek Luminometer. A
photomultiplier tube amplifies the
photons and results are reported as
Relative Light Units (RLU).Additional
identification is possible using the
residual broth left in the
Microfunnel.
Pall Corporation
GeneDisc Rapid
Microbiology
System
qPCR
Screening of
pathogens (e.g.
Specified
Microorganisms,
USP )
3 to 8 hours
including
sample
filtration,
nucleic acid
prep, and
PCR
96 per PCR
run
1-300 mL.Protocol
adapted for
solids and
non-filterable
samples.
1 CFU afterenrichment for 6
to 24 hours.
>100 CFU
without
enrichment.
E. coli, Salmonella,
Pseudomonas
aeruginosa,
Staphylococcus aureus,
Candida albicans,
Aspergillus brasiliensis,
STEC and non-STEC,
E. coli 0157, Listeria,
Legionella,
Enterococcus,
Cyanobacteria
Samples are filtered and
microorganisms collected on the
membrane are lysed by a combination
of reagents, sonication and heating.
The DNA and Master Mix (polymerase
and deoxynucleotides) are added to
the GeneDisc plate, pre-loaded with
the primers and probes. The plate is
transferred into the GeneDisc Cycler,
and it rotates through four
temperature zones during the PCR
amplification process. When the target
DNA sequence (from the
microorganism of interest) is
amplified, a fluorescent signal from
the probe will increase and is
measured in real time. Current plate
comes pre-loaded with 6 probes for
compendial specified microorganisms.
The GeneDisc Cycler can measure
fluorescent signal from each probe
simultaneously in real time resulting in
detection and positive identification of
microorganisms in a single PCR run of
less than 1 hour.
Roche
MycoTool
PCR
Mycoplasma
detection
< 5 hr
1 mL of cell
culture
containing 5
106cells/mL
< 1 CFU/mL> 90 species of
Mycoplasma
A 1 mL of sample of cell culture with a
concentration of 5 106cells/mL is
treated with lysis buffer. Mycoplasma
DNA is then purified and the target
DNA is amplified via PCR. The
resulting amplicons are evaluated
using gel electrophoresis.
Soleris
Neogen
Growth-based,
media based
detection via CO2or
pH
Quality, spoilage
and sterility
microbial testing
3 to 48 hours
Up to 128 per
unit; up to 4
units per
computer
0.1-5.0 mL1 CFU after
enrichment
Total aerobic count,
sterility, yeast & mold,
coliforms, E. coli, lactic
acid bacteria,
Enterobacteriaceae,
Pseudomonas,
Staphylococcus,
aciduric organisms
Samples are inoculated directly into a
vial which contains ready to use
media and a detection system. The
optical assay measures microbial
growth by monitoring pH or CO2that
generate a color change as
microorganisms proliferate.
Vivione
Biosciences, LLC
RAPID-B
Flow cytometry
Incoming inspection,
sterility confirmation,
bioburden, drug
20 min for
quantitative
results
Up to 8 hr for
96-160 per 8
hrs75-225 uL
1 CFU after
enrichment
TB, Chlamydia and
Staphylococcus for
clinical samples.
Salmonella, E. Coli
O157, non-O157
Samples and reagent are added to a
vial and mixed (5-10 min). The
RAPID-B system analyzes the sample
using flow cytometry, with results
available within 3-5 minutes. Light
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development,
process control
qualitative
results
STECs,
Staphylococcus, and
Vibrio for food samples
scatter resolution of around 130nm
shows separation of bacteria
populations via size and refractive
index alone. The system utilizes a
multi-parametric detection approach: a
phenotypic library to identify the
bacteria, an immunoprobe to
specifically tag the target organism
and a DNA dye to ascertain the live or
dead cells of the target organism. The
workflow is also based on the limit of
detection (LOD) required and thetarget of interest (may include
enrichment, filtration, centrifugation
and dilution).
RMMS FOR QUANTITATIVE ENUMERATION
Company
Product
Method
App lic ations
Time to
ResultThroughput
Sample Size
or TypeSensitivity
Organisms
DetectedWorkflow
Advencis
Lynx Technology
Growth based &
staining free;
High-Magnification
Imaging
Bioburden of raw
material, in-process
samples & finished
product, water
analysis
24-48 hr 240 per 48 hr
Filtrable
samples, solid
medium
1 CFU after
growth
Bacteria, yeast, mold,
spores
Samples are prepared according to
the compendial method (membrane
filtration or direct inoculation on solid
medium). The system incubates theplates (from 20 to 55C) and
automatically acquires high magnified
images (X50) of micro-colonies. The
image processing software analyzes
information on size and shape and
provides an enumeration for each
plate every 30 minutes. The system is
non-destructive, allowing for follow up
analysis.
AES Chemunex
D-Count and
BactiFlow
Viability staining and
flow cytometry
Bioburden of rawmaterial and
in-process samples,
finished product,
EM, water
30 min
150 per day
(BactiFlow)300 per day
(D-Count)
Liquids,
usually less
than 1 mL
10-50 cel ls Bacteria, yeast, mold
Viable cells in a liquid sample are
labeled with a non-fluorescent
substrate. Within the cytoplasm of
metabolically active cells, the
substrate is enzymatically cleaved (by
esterase) to release a fluorochrome.Cells with intact membranes will retain
the fluorescent label. The labeled
organisms pass through a argon ion
laser in the flow cell. Two fluorescence
detectors provide an enumeration in
cells per mL.
AES Chemunex
ScanRDI
Viability staining and
solid phase
cytometry
Bioburden of raw
material and
in-process samples,
finished product,
EM, water, sterility
testing
1.5-3 hr 30 per dayFilterable
samples1 cell
Bacteria, yeast, mold,
spores
The test sample is filtered through a
polyester membrane. Microorganisms
retained on the filter are labeled with a
non-fluorescent substrate. Within the
cytoplasm of metabolically active
cells, the substrate is enzymatically
cleaved (by esterase) to release a
fluorochrome. Cells with intact
membranes will retain the fluorescentlabel. An argon laser scans the
surface of the membrane within 3
minutes, and viable cells are detected.
Auto-fluorescent particles, membrane
fluorescence and background noise
are rejected and a total viable count is
reported. Viable cells may be
subsequently observed using a
phase-contrast microscope and an
automated stage.
Battelle
REBS
Raman
spectroscopy
Identification and
enumeration
3 min160 every 8
hr
Cells from
colony, liquid
medium,
product/raw
material,
surfaces
1 cell isidentified and
quantified
Alcaligenes,
Pseudomonas,
Brevundimonas,Candida, E. coli,
Bacillus, Ralstonia,
vegetative and spore
forms.
Sample material is retained on a
supported film. The area is examined
for microscopic particulates using
Raman spectroscopy and a spectral
signature is provided for eachparticulate. The spectral signatures
are statistically correlated to a library
of known microorganisms. No need
for Gram staining. Mixed cultures and
be identified and enumerated.
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Non-destructive for further analysis.
BD Diagnostics
FACSMicroCount
Viability staining and
flow cytometry
Fermentation
growth monitoring,
bioburden screening
of raw materials,
in-process samples,
finished product,
water analysis,
probiotics and stock
culture enumeration
4 min
quantitative;
24 hr for
enrichment
based
qualitative test
(presence /
absence)
12-15 per hr
for
quantitative
test;
20 samples
per hr for
qualitative
test
Up to 3 mL
30 to 106
quantitative;
1 cell qualitative
with enrichment
Bacteria, yeast, mold
For a quantitative test, the sample is
first diluted in buffer. For a qualitative
test (presence/absence testing), the
sample is diluted and inoculated into a
proprietary enrichment media, which
is then incubated for 24-48 hours.
Three mL of the diluted sample is then
loaded in the instrument for either test.
The automated system tags nucleic
acid of all microorganisms with
fluorescent stain. Sample is then
loaded into flow cytometer chamber
where a 639 nm red laser hits tagged
microorganisms to produce
fluorescence and side scatter signals.
Signals are captured for each
microorganism to reflect one count on
intensity plot. Total microbial count is
available in both tabular and graphical
formats.
BioVigilant
IMD-A
Light scattering
Active air monitoring
Instantaneous
Continuous
or episodic
monitoring
IMD-A 350
(28.3 L/min)
IMD-A 300
(1.15 L/min)
1 cellBacteria, yeast, mold,
spores
Air is drawn into the instrument.
Particles that pass through a 405 nm
diode laser are sized (0.5 to 10
microns) and enumerated using a Mie
scattering particle counter. At thesame time, particles that contain
biological targets, such as NADH,
riboflavin, and dipicolinic acid, will
auto-fluoresce as they pass through
the laser, and a separate fluorescence
detector will record these as viable
microorganisms. The system;
therefore, provides simultaneous
viable and total particulate data per
cubic volume of air. Results are
obtained in real-time, and there are no
consumables, reagents or media.
CCM MuScan
Innosieve
Diagnostics Test
Kits
Viability staining and
solid phase
cytometry;
fluorescence
microscopy
Bioburden of raw
material and
in-process samples,
finished product,
EM, water, sterilitytesting
10-65 minutes100 per 8
hours
Filterable
samples; 1 uL
to 1L
1 - 105cells
Bacteria, yeast, fungal
spores
The test sample is
filtered/concentrated through a
0.45um micro sieve. Microorganismsare retained on the membrane and
subsequently labeled with a viability
stain and/or a species-specific stain.
Available are total viable count, total
live-dead ratio, total species-specific
count and total species-specific viable
count. After staining a scanning period
using MuScan digital fluorescent
microscopy at specific excitation and
emission wavelengths is performed.
The image processing software
analyzes fluorescent objects on size,
shape and fluorescent signals of all
microbes. The assay can be
user-customized with different types of
fluorescent stains, labeled antibodies,
DNA-probes, etc., depending on the
organism(s) to be detected. Examples
of test kits for specific organisms
include Legionella, Salmonella,
Listeria, Chronobacter and E. coli. The
method is non-destructive such that
detected microbes can be
subsequently cultured.
Lonza
MicroCompass II
Note that this
RMM is currently
being redesigned
and is currently
unavailable.
RT-PCR
Estimation of cell
count
4-5 hr 72 per 8 hr ~ 1 mL 100 cells Bacteria, yeast, mold
Microorganisms are captured from
filterable samples via spin filtration,
and are then chemically lysed to
release nucleic acid. Alternate
procedures are used for non-filterable
samples. Nucleic acid extraction and
purification is fully automated using
magnetic beads and washing steps.
Purified DNA is amplified using PCR
and ribosomal RNA is amplified using
reverse transcriptase (RT) PCR. PCR
is monitored in real-time (via a MGB
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Eclipse probe), and the number of
cycles where the fluorescence rate
increases above a threshold value is
used to estimate the number of viable
cells in the original sample.
Appropriate studies are required to
correlate the number of PCR cycles
with cell concentration.
Millipore
Milliflex Rapid
Growth-based; ATP
bioluminescence
Bioburden of raw
material and
in-process samples,
finished product,
EM, water, sterility
24-28 hr 60 per dayFilterable
samples
1 CFU after
growthBacteria, yeast, mold
Utilizes a membrane filter to capture
individual cells. Filter the sample and
place the membrane on an
appropriate agar medium to allow the
growth of micro-colonies. Micro-
colonies are then treated with an
ATP-releasing reagent, followed by
the addition of luciferin and luciferase.
Photons of light from each micro-
colony are detected by a luminometer,
and a cell count is reported. May be
able to continue incubation to form
larger colonies for subsequent
microbial identification.
Millipore
Milliflex Quantum
Growth-based;
viability staining and
cellular fluorescence
Bioburden of raw
material and
in-process samples,
finished product,
EM, water, sterility
24-28 hr 60 per dayFilterable
samples
1 CFU after
growthBacteria, yeast, mold
Filter the sample, and incubate the
membrane an an agar cassette to
form micro-colonies. Add
non-fluorescent substrate andincubate an additional 30 min. Within
the cell, the substrate is enzymatically
cleaved, releasing a free fluorochrome
into the microorganism cytoplasm. As
fluorochrome accumulates inside the
cells, the signal is naturally amplified.
The membrane is placed in a reader
and exposed to the excitation
wavelength of the fluorochrome.
Fluorescent micro-colonies are then
automatically enumerated.
Non-destructive; can continue to
incubate media to obtain colonies for
microbial identification.
Pall Corporation
GeneDisc Rapid
MicrobiologySystem
qPCR
Estimation of cell
count
3 to 8 hours
including
sample
filtration,
nucleic acidprep, and
PCR
96 per PCR
run
1-300 mL.
Protocol
adapted for
solids and
non-filterable
samples.
1 CFU after
enrichment for 6
to 24 hours.
>100 CFU
without
enrichment.
E. coli, Salmonella,
Pseudomonas
aeruginosa,
Staphylococcus aureus,
Candida albicans,
Aspergillus brasiliensis,
STEC, E. coli 0157,
Listeria, Legionella,
Enterococcus,
Cyanobacteria
Samples are filtered andmicroorganisms collected on the
membrane are lysed by a combination
of reagents, sonication and heating.
DNA is purified and concentrated by
further filtration. The DNA and Master
Mix (polymerase and
deoxynucleotides) are added to the
GeneDisc plate, preloaded with the
primers and probes. The plate is
transferred into the GeneDisc Cycler,
and it rotates through four
temperature zones during the qPCR
amplification process. With each
amplification cycle a fluorescent signal
is generated from the probe. The point
at which the signal reaches above the
background is called Cycle Threshold
(Ct) value. The higher the DNA copy
(e.g., higher microorganisms), the less
amplification cycles will be required to
reach the Ct value (i.e., faster
detection). Built-in software calculates
the number of genomic copies (e.g.,
amount of DNA, which is directly
related to the number of microbial
cells/CFUs) present in the initial
sample. Identification is possible by
DNA sequencing of the remaining
DNA sample.
Particle
Measuring
Systems
BioLaz
Real-Time
Microbial Monitor
Light scattering
Active air monitoring
InstantaneousContinuous
monitoring3.6 L/min 1 cell
Bacteria, yeast, mold,
spores
Air is drawn into the instrument
sensing area via a stainless steel
sample probe. As the air passes
through the system it is illuminated by
a laser. Biological particles that
contain NADH or riboflavin will
auto-fluoresce as they pass through
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the laser. Those fluorescing biological
particles are then counted in one of
two size channels. The system can be
integrated into an existing
Environmental Monitoring data
management platform or can be used
with a local PC with available interface
software.
rap.ID Particle
Systems
rap.ID
Viable Staining and
Imaging LED
Raman
Spectroscopy
Identification and
enumeration
3-10 min300-600 ID's
per hour
Cells from
colony, liquid
medium,
product/raw
material,
surfaces
1 cell is
identified and
quantified
> 150 bacterial and
spore entries;
customizable
The sample material is collected on
metal foil using impaction or filtration
methods. Viability staining and
automated image analysis using dark
field illumination detects viable particle
quantity, shape, and size ranging from
0.5 m and larger. Raman
spectroscopy is then performed on
each viable particle and a spectral
signature is provided. The spectral
signatures are statistically correlated
to a library of known microorganisms.
Non-destructive for further analysis.
Rapid Micro
Biosystems
The Growth
Direct System
Growth-based;
automated detection
of cellular
auto-florescence
Water analysis,
Bioburden, Sterility,
Environmental
Monitoring
Final results in
one-half the
time of the
compendial
method
Positive
results within
hours
400 samples
for 3-day EM
Test
280 samples
for 5-day
water or
bioburden
tests
20-40
samples per
day for
Sterility Test
Filterable
samples
Standard
Environmental
Monitoring
samples
1 CFU
Bacteria, yeast, mold,
all organisms that grow
on agar media
Samples are prepared as per the
compendial method and loaded into
the system which manages the
incubation and colony enumeration for
the sample. Proprietary digital imagingtechnology automatically enumerates
micro-colonies in one-half the time
than traditional visual plate counting
methods. The sample is collected onto
a filter placed onto an agar medium
cassette with an optically clear lid.
Illumination with blue light excites
micro-colonies to auto-fluoresce
(without the addition of any reagents),
which are enumerated by a CCD
imaging system. The system
automatically incubates and analyzes
each cassette for the formation of
micro-colonies over time. Particles
that do not grow in size over time are
ignored. Non-destructive; can
continue to incubate media to obtain
colonies for microbial identification.
TSI Inc.
BioTrak
Real-Time Viable
Particle Counter
Light scattering
Active air monitoring
Instantaneous
Continuous
or episodic
monitoring
28.3 L/min 1 cellBacteria, yeast, mold,
spores
Air is drawn into the instrument and
both viable and total particulates are
simultaneously detected, sized and
enumerated via a 685 nm laser diode
(for particle sizing) and a 405 nm laser
diode (for viability detection). The size
range for detection is from 0.5 to 25
microns. The counting efficiency is
50% at 0.5 microns and 100% for
particles > 0.75 microns (per ISO
21501-4 and JIS B9921). The system
can be calibrated using NIST
traceable standards. Also included is
an integrated particle collection filter
(gelatin filter) to capture
microorganisms for subsequent
growth and identification.
Vivione
Biosciences, LLC
RAPID-B
Flow cytometry
Incoming inspection,
sterility confirmation,
bioburden, drug
development,
process control
20 min for
quantitative
results
Up to 8 hr for
qualitative
results
96-160 per 8
hrs75-225 uL
1 CFU after
enrichment
TB, Chlamydia and
Staphylococcus for
clinical samples.
Salmonella, E. Coli
O157, non-O157
STECs,
Staphylococcus, and
Vibrio for food samples
Samples and reagent are added to a
vial and mixed (5-10 min). The
RAPID-B system analyzes the sample
using flow cytometry, with results
available within 3-5 minutes. Light
scatter resolution of around 130nm
shows separation of bacteria
populations via size and refractive
index alone. The system utilizes a
multi-parametric detection approach: a
phenotypic library to identify the
bacteria, an immunoprobe to
specifically tag the target organism
and a DNA dye to ascertain the live or
dead cells of the target organism. The
workflow is also based on the limit of
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k
detection (LOD) required and the
target of interest (may include
enrichment, filtration, centrifugation
and dilution).
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