Data SheetData Sheet
Guava® ViaCount® Assay Rapid, accurate cell counting and viability assessments
The Guava ViaCount Assay is fast becoming the new
standard for viability and cell counting. In this simple
no-wash, mix-and-read-assay, you can accurately
obtain absolute total cell counts, perform viability
assessments, and determine apoptotic percentages—
all from tiny samples.
You’ll enjoy several advantages over traditional
methods, including greater accuracy, reproducibility,
and speed. This assay eliminates the user-to-user
variability characteristic of trypan blue hemocytometer
counting. As a result, you have significant increases in
counting reproducibility. In addition, the Guava ViaCount
Assay is more accurate than hemocytometer counting
or impedance-based counting over a wide range of
cell concentrations. Cell counting with this assay has
also proven to be up to 10 times faster than manual
hemocytometer methods. Unlike traditional cytometer
counting, you don’t need to use expensive reference
beads because of Guava’s unique capillary technology.
Cell counting and viability assessments can be used
in a variety of applications, such as cytotoxicity studies,
PBMC counting, rapid apoptosis assessment, and
increasing reproducibility in cell-based assays.
FeaturesAssay:
• Simple no-wash, mix-and-read procedure
• Analyzes viability of individual cells
• Delivers direct, absolute cell counts and population
percentage
• Counts up to 10 times faster than manual methods
• More reproducible than traditional tests
Samples: • Uses small samples in tubes or a 96-well plate
• Handles low-density and small-volume cell samples
• Works with adherent or suspended cells and mammalian
and insect cells
Compatibility: • Runs on Guava EasyCyte™ Mini, EasyCyte Plus, PCA,
and PCA-96 Systems
Debris Viable Cells Apoptotic Dead Cells
Forward Low High High HighScatter
Nuclear Neg High High HighStain
Viability Neg Neg Med HighStain
2
Mean
1 stdv
2 stdv
3 stdv
1200
800
600
2000
1800
1400
1000
1600
cel
ls6
SFC
/10
Assay Number
1 2 3 4 5 6 7 8
Manual via Trypan Blue
Guava ViaCount
Figure 3. Increased reproducibility in ELISPOT assays when using
Guava ViaCount assay for cell counting.
Figure 1. Guava ViaCount assay provides accurate cell counts.
Comparisons with an impedance counter and manual trypan blue
hemocytometer counting indicated greater linearity and accuracy over a
range of cell concentrations. Expected cell counts were determined using
an FDA-cleared reference counting bead method 1.
cel
ls/m
L) 8
6
6O
bser
ved
Cou
nt (x
10
4
2
0
Expected Count (x106 cells/mL)
0 2 4 6
Hemacytometer Impedance Countery=0.95x + 0.23 y=1.27x + 0.17R2=0.877 R2=0.964
Guava PCAy=0.94x + 0.25R2=0.966
Guava PCA ( )
Hemacytometer ( )
Impedance Counter ( )
HOW THE GUAVA VIACOUNT ASSAY WORKSThe ViaCount assay uses a proprietary mixture of two
DNA binding dyes to provide sensitive, accurate
detection of viable, apoptotic, and dead cells.
A membrane-permeant dye stains all nucleated
cells, leaving cellular debris unstained so that you can
focus on whole cells. A membrane-impermeant dye
stains only damaged cells (i.e. those with breaches in
the plasma membrane), thus indicating apoptotic and
dying cells.
QUICKLY IDENTIFY APOPTOTIC CELLS With the Guava ViaCount assay, you can easily identify
apoptotic cells using the viability dye (figure on front
and figure 2). You can also use an optional apoptosis
gate in the analysis software that counts apoptotic
cells separately. This optional gate feature enables you
to use the assay for cytotoxicity, apoptosis induction,
or proliferation studies. In these cases, more than a
mere cell number is essential to understanding an
experiment’s outcome.
By contrast, published studies show that trypan
blue exclusion assays often overestimate cell counts2,3.
This may occur because trypan blue cannot identify
apoptotic cells, and so they are added to the count of
viable cells.
Figure 2. The blue population of cells show a significant amount of
annexin V staining (bottom plot), indicating that intermediate levels of
staining with the viability dye correlates with apoptosis.
Nuc
leat
ed C
ells
Viability Stain
Live
Dead
Apoptotic Via
bilit
y Sta
in
Annexin V
Dead
Live
Apoptotic
IMPROVE YOUR CELL-BASED ASSAYSAccurate cell counts and viability measurements make
other functional cellular assays more reproducible
because they provide a more realistic indication of how
many healthy cells are present. For example, ELISPOT
assays show a marked reduction in CVs when cell
concentrations are measured with the Guava ViaCount
assay compared to those done with trypan blue
hemocytometer measurements (Figure 3).
COMPATIBLE WITH MANY CELL TYPESThe Guava ViaCount assay accurately determines
viability and counts for a large variety of cell types.
Tested on a broad selection of mammalian cell lines
(including human, mouse, rat, hamster, and monkey). As
a plus, the assay also works with both primary cells and
whole blood.
3
EASY-TO-USE SOFTWARE Guava’s ViaCount Software Module makes data
collection and analysis even easier. It runs the entire
assay without any need for user input, eliminating
subjectivity. Sample analysis is completed and the data
are automatically exported to a time-stamped
spreadsheet in seconds. Plus, Guava’s new EasyFit
Software is included for computer-aided population
analysis.
The module displays data in user-friendly formats
so you can monitor your experiment or read data right
off the screen. By measuring the staining intensities of
the two dyes, our software represents each cell
counted as a dot on the Cell Dotplot. Cell counts and
percentages are calculated and displayed real-time on
the screen. The optional apoptosis gate (purple line)
can easily be unchecked if only the viable population is
relevant to your research.
Benefits:• Easier to learn and use • Get standardized results even with multiple users • Spend less time optimizing instrument settings • Improve your productivity
ADDITIONAL REAGENTS FOR GREATER FLEXIBILITYCreated to address more difficult cell counting
samples, Guava ViaCount Flex reagent is a more
concentrated formulation of the ViaCount reagent. It
works well for low density cell cultures since it can be
added in small volumes to avoid further sample dilution.
This feature also makes ViaCount Flex reagent ideal for
cell counting and viability assessment in 96-well plates.
For clumped samples, use the ViacCount Cell
Dispersal Reagent, which uses enzymes to gently
disaggretate cells. This helps to improve the accuracy
and precision of cell counts.
ViaCount Flex Reagent• Low density cell samples that are too diluted to be
counted with standard hemacytometers• Cell lines that stain heterogeneously (facilitates gate
settings by providing tighter staining patterns)
Viacount Flex Reagent for Challenging Samples• Non-mammalian cell lines (such as SF9 insect cells)
and other cell lines with unusual culture conditions
Guava ViaCount Cell Dispersal Reagent (CDR) • To reduce clumping in adherent cells
A. Easy viewing of
real-time statistics
for cell counts and
concentrations of
original sample.
B. Optional apoptosis
gate (purple line)
identifies apoptotic
cells (intermediate level
staining, red circle).
C. Real-time counts of
cells in all populations
(apoptotic, dead, viable)
with calculated cell
concentrations.
A
2B 1BC
ORDERING INFORMATIONDescription Catalogue No.
Kits & ReagentsGuava ViaCount Reagent Kit (100 tests) 4000-0040
Guava ViaCount Reagent Kit (600 tests) 4000-0041
Guava ViaCount Flex Reagent for Challenging Samples (100 tests) 4500-0110
Guava ViaCount Cell Dispersal Reagent (100 tests) 4700-0050
Guava ViaCount Flex Reagent (500 tests) 4700-0060
SoftwareViaCount Software, PCA System 0500-1030
ViaCount Software, PCA-96 System 0500-0580
ViaCount Software, EasyCyte MiniSystem 0500-1470
ViaCount Software, EasyCyte Plus System 0500-2000
www.millipore.com
Millipore is a registered trademark of Millipore Corporation. The M mark is a trademark of Millipore Corporation. Guava, the Guava logo, Guava ViaCount, and Guava Technologies are registered trademarks of Guava Technologies, Inc. EasyCyte is a trademark of Guava Technologies, Inc.Lit. No. DS0125EN00 07/08 Printed in U.S.A. BS-GEN-08-00631 © 2008 Millipore Corporation, Billerica, MA 01821 U.S.A. All rights reserved.
Audience
GuavaAssays & Kits
Flow Cytometry Systems Service
Europe & North America
Academic & Government n n n
Pharma & Biotech l l n
All Other Countries n n n
n Millipore Corporation l Guava Technologies
To learn more about our integrated solutions for
flow cytometry, please contact Millipore or Guava
Technologies. The chart (below) shows which company
serves your organization.
To stay up-to-date on our joint product and
service offerings, please visit
www.millipore.com/flowcytometry.
www.guavatechnologies.com
References
1. Data provided courtesy of Amy Hirst, Raven Biotechnologies, Inc.
2. Mascotti, McCullogh and Burger; Transfusion 40:693-696, 2000.
3. Altman, Rander and Rao; Biotechnology Progress 9:671-674, 1993.
4. Ogasawara K, et al. Impairment of NK cell function by NKG2D modulation in NOD mice. Immunity. 18(1): 41-51 (2003).
5. Arase H, et al. Direct recognition of cytomegalovirus by activating and inhibitory NK cell receptors. Science. 296(5571):1323-1326 (2002).
6. Cerwenka A, et al. Cutting edge: the minor histocompatibility antigen H60 peptide interacts with both H-2Kb and NKG2D. Journal of Immunology. 168(7):3131-3134 (2002).
7. Ogasawara K, et al. Inducible costimulator costimulates cytotoxic activity and IFN-gamma production in activated murine NK cells. Journal of Immunology. 169(7):3676-3685 (2002).
8. Phi-Wilson J, et al. A technology for the rapid acquisition of cell number and viability. American Biotechnology Laboratory. pp 34-36 (2001).