Cellometer K2 Fluorescent Viability Cell Counter

Fluorescent automated cell counter for complex primary samples


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The Cellometer K2 is fast and very accurate. We love that the dye is nuclear to ensure accuracy of the cell counts. Sure beats counting by hand!

Amanda Clark, Biomere

The Cellometer K2 is great and easy to use. It gives clear images quickly. We have been using it for a couple years and never have any problems with it!

Solaema Taleb, IUPUI

The Cellometer K2 is very fast and efficient. It has turned what used to be a hassle in the lab into a breeze!

Andrew Schade, KCAS

Cellometer K2 is very compact, reliable and easy to use. Our lab uses this counter to count live and dead cells for our experiments. It is a very good alternative to flow cytometer for cell cycle and apoptosis studies.

Stanford University

We really like this instrument [Cellometer K2]. We were counting cells manually until now. This has made our life much easier.

MD Anderson Cancer Center

Features of the Cellometer K2 Image Cytometry System

Simple, Automated Cell Counting in 60 Seconds

The Cellometer K2 utilizes bright field imaging and dual-fluorescence imaging to quickly and accurately identify and count individual cells. Cell count, concentration, diameter, and % viability are automatically calculated and reported.

Results in less than 30 seconds Load Sample, View Image, Count Cells, and Obtain Results in < 60 seconds

GMP/GLP Support Module Available for Access and Audit Control

  • Four different access level controls for user logging
  • Audit trail by user
  • Autolocking of system (inactivity timeout)
  • Password aging reset requirement
  • Auto printing option
K2 software feature screenshot

Simple, User-friendly Procedure

Simple, User-friendly Procedure - pipette load sample for Cellometer K2, cell viability counter

Pipette 20µl Insert Slide Select Assay & Click Count View Results in 60 seconds!

The K2 Allows Users to:

  • Increase throughput
  • Increase accuracy
  • Improve consistency
  • Ensure all data is correctly captured
  • Count difficult cells (clumpy, irregular-shaped)
  • Eliminate judgment errors, miscounts, interference from red blood cells and user-to-user variability

Primary Cell Analysis: PBMCs, Hepatocytes, and more

The Cellometer K2 is specifically optimized for analysis of primary cells from peripheral blood, cord blood, bone marrow, and other complex samples for use in a wide range of research areas, including:

  • Nucleated Cells for Transplantation
  • PBMCs for Immunology
  • Splenocytes for Vaccine Development
  • Stem Cells for Cellular Therapy
  • Tumor Cell Suspensions for Oncology

Dual-color fluorescence allows for staining of live and dead nucleated cells, generating accurate viability results even in the presence of debris, platelets, and red blood cells. Accurate analysis of both ‘messy’ and ‘clean’ samples enables the K2 to evaluate samples at a variety of points throughout sample processing – from initial collection to separation, to cryopreservation.

The Cellometer K2 features assays for analysis of a wide range of primary samples, including:

Hepatocytes - Cellometer K2, cell viability counter

Hepatocytes

Jurkat - Cellometer K2, cell viability counter

Jurkat

Splenocytes - Cellometer K2, cell viability counter

Splenocytes

PBMC - Cellometer K2, cell viability counter

PBMC

Live / Dead Nucleated Cell Counts using Dual-Fluorescence

Green fluorescent live cell image

Green fluorescent live cell image - Cellometer K2, cell viability counter

Red fluorescent dead cell image

Red fluorescent dead cell image - Cellometer K2, cell viability counter

Why Dual-Fluorescence?

Because bright field cell counting does not differentiate nucleated from non-nucleated cells and trypan blue staining is not as easy to detect as fluorescent staining, dual-color fluorescence is strongly recommended for accurate viability analysis for primary cells. The K2 is equipped with standard assays for dual-fluorescence analysis of primary cells stained with AO/PI.

The AO/PI Method

Acridine Orange, AO, is a nuclear staining (nucleic acid binding) dye permeable to both live and dead cells. It stains all nucleated cells to generate green fluorescence. Propidium iodide, PI, can only enter dead cells with compromised membranes. It stains all dead nucleated cells to generate red fluorescence. Cells stained with both AO and PI fluoresce red due to quenching, so all live nucleated cells fluoresce green and all dead nucleated cells fluoresce red.

Cell-Based Assays: Cell Cycle, Apoptosis and GFP

Cell Cycle

Cell Cycle - Cellometer K2, cell viability counter

Cellometer K2 Image Cytometer has the ability to perform basic cell-based assays such as cell cycle, apoptosis, and green fluorescent protein (GFP) population analysis. These cell-based assays can be performed by exporting image cytometric analysis data to FCS Express from De Novo Software for data analysis and presentation.

For cell cycle analysis, the different cell cycle phases can be analyzed using the cell cycle kit from Nexcelom Bioscience to determine the SubG1, G0/G1, S, and G2/M phase cell population. For apoptosis analysis, using Annexin V-FITC/PI and Caspase 3/8 staining Kit from Nexcelom Bioscience to determine percent apoptotic cell population. GFP expression percent population can also be directly measured using Cellometer K2.

Apoptosis

Apoptosis Negative Control - Cellometer K2, cell viability counter

Untreated (Negative Control)

Healthy Apoptotic Necrotic Debris
AVE 81.9% 8.1% 4.0% 6.0%
STD 1.6% 1.3% 0.4% 1.1%
CV 1.9% 16.1% 9.3% 18.3%

Apoptosis Positive Control - Cellometer K2, cell viability counter

Treated (Positive Control)

Healthy Apoptotic Necrotic Debris
AVE 58.8% 24.7% 13.8% 2.7%
STD 1.9% 1.1% 1.2% 0.2%
CV 3.2% 4.3% 9.0% 9.2%

No Interference from Red Blood Cells, Platelets, or Debris

The dual-fluorescence AO/PI method utilizes nuclear staining dyes that bind to nucleic acids in the cell nucleus. Because most mature mammalian red blood cells do not contain nuclei, only live and dead mononuclear cells produce a fluorescent signal. There is no need to lyse red blood cells, saving time and eliminating an extra sample preparation step. Red blood cells, platelets, and debris are not counted in the fluorescent channels.

The advantage of fluorescent counting for primary cells

These images (right) demonstrate the advantage of fluorescent counting for primary cells. The bright field image shows the combination of nucleated cells, red blood cells, and platelets present in the sample. Only the live and dead nucleated cells are visualized and counted in the green and red fluorescent channels.

Sample Measurement Total nucleated All RBC % RBC n
PBMC+RBC Mean

CV

1.26E+07

6.2%

1.39E+07

8.8%

1.23E+06 8.9% 10
PBMC+1/2RBC Mean

CV

1.21E+07

4.8%

1.27E+07

5.4%

5.82E+05 4.6% 10
PBMC+1/4RBC Mean

CV

1.22E+07

7.9%

1.24E+07

7.3%

2.27E+05 1.8% 10

Fresh human PBMCs (peripheral blood mononuclear cells) were spiked with varying amounts of RBCs (red blood cells.) All cells (nucleated + RBC) were counted in the brightfield channel. Nucleated cells were then counted in the green fluorescent channel. Varying amounts of RBCs (1.8%, 4.6%, and 8.9%) did not affect the nucleated cell count.

Fresh human PBMCs - Cellometer K2, cell viability counter

Several red blood cells are indicated in the bright field image (above, left). The red blood cells are not visible in the fluorescent image (above, right) detecting cells stained with nuclear staining dye.

Cell Images for Data Verification

No two cells are the same.

With the Cellometer K2 Image Cytometer, cell morphology can be immediately viewed on-screen in the bright field image.

Counted cells are indicated on-screen for further verification that cells in the sample are being imaged and analyzed properly. Bright field counted images can be viewed for basic cell counting and trypan blue viability.

Fluorescent counted images indicating counted live and dead nucleated cells can be viewed for dual-fluorescence primary cell viability assays.

Users can confirm that:

  • cells are counted correctly, based on size and shape
  • cells within clumps are being counted individually
  • red blood cells, platelets, and debris are being excluded from results

Bright field counted cell image for Data Verification - Cellometer K2, cell viability counter

Fluorescent counted cell image for Data Verification - Cellometer K2, cell viability counter

The bright field image confirms that individual cells within pairs are being counted and smaller debris is not being counted. In the combined fluorescent counted image, live counted cells are circled in green. Dead counted cells are circled in red.

  • Cell images can be archived and exported for use in publications and presentations.
  • Saved images can be re-counted using default or user-optimized analysis settings

Cellometer Primary Hepatocyte Viability Analysis Method

Due to hepatocytes’ variable morphology, fragile nature, and tendency to clump, traditional manual counting methods can be time-consuming and inaccurate. Because hepatocytes lose viability over time, extended or variable counting times can generate inaccurate and inconsistent viability determinations. Hepatocytes are also too fragile to evaluate using flow cytometry due to flow-induced shear stress. Cellometer image cytometry is the most reliable method for determination of hepatocyte viability.

Dual-fluorescence Staining Procedure

For viability determination, 20µl of hepatocyte sample is mixed with 20µl of Cellometer AO/PI Staining Solution. The acridine orange (AO) dye stains DNA in all nucleated cells, generating green fluorescence and easily differentiating hepatocytes from debris. Propidium iodide (PI) stains DNA in all cells with compromised cell membranes, generating red fluorescence. In cells stained with both AO and PI, the green fluorescence is absorbed by the red fluorescence via FRET (fluorescence resonance energy transfer), so all dead hepatocytes fluoresce red and can be easily counted. The procedure is fast, gentle, and accurate.

Dual-fluorescence Staining Procedure - Cellometer K2, cell viability counter

Bright field image (left) shows the variable morphology of primary hepatocytes. Dual fluorescence image (right) shows counted live hepatocytes (circled in green) and counted dead hepatocytes (circled in red).

Cellometer Analysis - Cellometer K2, cell viability counter

Cellometer Analysis

Immediately after mixing, 20µl of stained sample is loaded into the Cellometer Counting Chamber and inserted into the Cellometer K2 instrument. The sample is imaged directly from the counting chamber. Because the counting chamber is disposable, no washing is required between samples and there is no risk of cross-contamination. Samples are imaged and analyzed using pre-set parameters for primary hepatocytes.

Analysis of Clumpy and Irregular-shaped Cells

Including NCI-60 and clumpy MCF-7 Cells

NCI-60 is a group of 59 human cancer cell lines (originally 60) developed by the National Cancer Institute for screening purposes.

  • 57% of the NCI-60 cell lines are clumpy, contain debris, or display large variations in cell shape or size
  • All 59 NCI-60 cell lines have been successfully validated on the Cellometer Image Cytometer

All 40 of the NCI Comprehensive Cancer Centers use Cellometer Cell Counters.

Clumpy Cells - Cellometer K2, cell viability counter

Clumpy Cells

The MCF-7 breast cancer cell line can be very clumpy. The Cellometer pattern-recognition software identifies and counts individual cells within these cell clumps for accurate analysis (shown above).

Irregular-shaped Cells - Cellometer K2, cell viability counter

Irregular-shaped Cells

The Cellometer cell roundness setting can be adjusted for recognition and counting of irregular-shaped cells, such as RD cells and activated T-cells.

Cell Size Analysis & Size-based Counting

PBMC cell diameter histogram

The Cellometer K2 Image Cytometer Automatically generates a cell size histogram based on cell diameter.

Because Cellometer generates individual cell size measurements, multiple samples can be overlaid on one histogram enabling analysis of the change in cell diameter over time.

10x Faster than Manual Counting

Counting 1 x 106 cells takes approximately 5 minutes with a manual hemacytometer. Counting live and dead cells sometimes takes twice as long. The Cellometer K2 Image Cytometer calculates cell count and concentration for live and dead cells and % viability in just 60 seconds.

Hemacytometer under microscope using Cellometer K2, cell viability counter

Improve Data Accuracy & Consistency

  • Eliminate Wash Steps
  • Eliminate Judgment Errors
  • Eliminate interference from RBCs
  • Eliminate Recording & Calculation Errors
  • Reduce Counting Time … Run More Experiments

Cellometer Precision

The Cellometer K2 Image Cytometer offers excellent reproducibility, with a %CV (Coefficient of Variation) of <10% for fluorescent concentration and viability measurements. The data (right) is based on four preparations of Jurkat cells stained with propidium iodide, a fluorescent nuclear-staining dye.

Sample N Value Average Live Cell Concentration % Viability CV of Concentration CV of Viability
Jurkat 24 3.61E+06 92.2% 8.9% 1.0%
Human PBMC 10 5.94E+06 96.0% 4.7% 0.5%
Mouse Splenocyte 10 1.86E+07 88.6% 5.6% 0.7%

Imaging / Counting Chambers: No Washing or Contamination

Cellometer disposable images chambers for Cellometer K2, cell viability counter
Cellometer Disposable Imaging Chambers consist of two independent enclosed chambers with a precisely controlled height. Cell suspension of 20 microliters is loaded into the chamber using a standard single channel pipette. The chamber is inserted into the Cellometer cell counter and the cells are imaged. This simple sample loading and analysis method is ideal for fragile cells.

The disposable Cellometer Cell Counting Chambers offer several key advantages:

  • Time savings – no washing
  • No risk of cross-contamination
  • Reduced biohazard risk to users
  • Controlled sample volume
  • Large-depth chambers for large cells

Applications for Cellometer K2 Fluorescent Viability Cell Counter

Adipocytes

Adipocytes

Automatically measure cell size of freshly isolated adipocytes and plot size histogram. DNA staining fluorescence dyes are used to identify cells from lipid droplets. »

Adoptive Cell Transfer Therapy

Adoptive Cell Transfer Therapy

Use Cellometer to perform cell based assays and measure cell size, viability and concentration of cell lines and primary samples used in adoptive cell therapy research. »

Apoptosis

Apoptosis

Automatically detect and analyze Caspase3 and 8, JC-1, and Annexin V apoptotic events using the Cellometer image cytometery. »

Cell Cycle

Image Cytometry for Cell Cycle Analysis

Automatically measure the cell cycle of mammalian cells. Generated cell cycle histogram allows for easy data analysis and presentation. »

Trypan Blue and AO/PI

Cell Viability Measurement Using Trypan Blue or AO/PI

When should you use trypan blue and when should you use acridine orange/propidium iodide to measure cell viability? »

Cell Viability and Necrosis

Cell Viability and Necrosis

Cell viability is performed using various fluorescent membrane exclusion dyes, such as PI, EB, 7AAD, and others. This assay is performed by enumerating cells in captured bright-field and fluorescent images. And Necrotic cells are detected using propidium iodide. »

Blood Samples

TNC Concentration & Viability for Clinical (Blood) Samples

Analyze fresh and processed blood and bone marrow samples without lysing: no interference from RBCs. »

Cell Size Assay

Cell Size Assay

Performing cell size measurement assay and using cell size to count cells within preset cell size parameters. For adipocytes, stem cells, Sf9 cells, dendritic cells, and others. »

Cancer Cell Lines

NCI-60 Cancer Cell Lines

Automatically measure live cell concentration and viability of cancer cell lines used in oncology research and most of all biology research. »

GFP Transfection Efficiency

Quantitative Measurement of GFP Transfection

Rapidly identify fluorescence positive cells from a sample, analyze individual cell fluorescence intensity, calculate cell concentration, size and determine the GFP transfection automatically. »

Hepatocytes

Fresh & Cryo Preserved Primary Hepatocytes

Automatically measure live hepatocyte concentration and viability using dual fluorescent nuclear stains, for human, rat, mouse and horse. »

Immunology

Immunology Research

Automatically quantify cell viability and concentration for a variety of immunologically relevant samples such as: bone marrow, cord blood, slpenocytes, lymphocytes, isolated mononuclear cells, tumor digests, murine samples, and others. »

Insect Cells

Insect Cells

Automatically measure live cell concentration, viability for baculovirus infected insect cells. Cell size histogram live cell concentration and viability are generated within less than 60 seconds using 20 µl sample. »

PBMC

Peripheral Blood Mononuclear Cells (PBMC)

Automatically measure live cell concentration and viability without lysing red blood cells for consistent results from patient samples. Other cells include splenocytes and bone marrow. »

WBSc in Whole Blood

WBCs in Whole Blood

Automatically measure nucleated cell concentration without lysing red blood cells using nuclear staining dyes (AO), for human and mouse blood. »

Performance of the Cellometer K2 Image Cytometer

Total Cell Concentration Range of Jurkat Cells Measured by Cellometer K2

Concentration Dynamic Range Figure 1 depicts the dynamic range for cell concentration measurements on Cellometer K2. This data set was taken on a concentration series of cultured Jurkat cell line.

Samples from 1 x 105 – 1 x 107 cells/ml can be counted without further dilution.

The %CV at each concentration was below 10%.

Dynamic Range of Total Cells with K2

Figure 1: Table of results for cell concentration dynamic range

Cellometer K2 Repeatability and Consistency

Sample N Value Average Live Cell Concentration % Viability CV of Concentration CV of Viability
Jurkat 24 3.61E+06 92.2% 8.9% 1.0%
Human PBMC 10 5.94E+06 96.0% 4.7% 0.5%
Mouse Splenocyte 10 1.86E+07 88.6% 5.6% 0.7%
Figure 2: Table of results for cell concentration and viability using AO/PI

The results indicate the accuracy of the Cellometer K2 instrument in assessing the viability of Jurkat cells using AO/PI for cell viability. Jurkat, human PBMC, mouse splenocytes were tested at 24, 10, and 10 sample replications, respectively. The viability average was calculated and plotted. The results show the reliability and accuracy of the Cellometer K2 in measuring cell concentration and viability of mammalian cells.

Consistency and Accuracy Comparison to Hemacytometer

Jurkat

N = 20 Hemacytometer Cellometer K2
Average 1.03E+06 1.04E+06
STDEV 6.60E+04 5.57E+04
%CV 6.4% 5.3%

5 µm beads

N = 20 Hemacytometer Cellometer K2
Average 1.07E+06 1.03E+06
STDEV 5.90E+04 5.38E+04
%CV 5.5% 5.2%

Cellometer K2 Specifications

Includes
  • Cellometer K2 Instrument
  • Cellometer Software
  • Two Standard Fluorescence Optics Module
  • Power Supply
  • USB 2.0 Connection Cable
  • Phone/online applications support during set-up
Available Accessories
  • PC Laptop
Imaging Performance Cell Size 5 – 300* microns
Conc. Range: 105 – 107 cells/mlBright field imaging, fluorescent imaging and pattern-recognition software to quickly and accurately decluster, identify and count individual cells.* Cellometer CHT4-PD300 Slides are required for cells > 80 microns in diameter
Instrument Specifications Weight: 23.0 lbs. (10.4 kg)
Dimensions: Width: 6.0” (15.2 cm), Depth: 8.5” (21.6 cm), Height: 14.0” (35.6 cm)
Input to Power Adapter: 100-240 VAC, 50/60 Hz, 1.0A
Output to Instrument: 12 VDC, 3.34A
PC / Laptop Minimum Requirements: (If purchasing Cellometer without PC laptop)
  • Windows 7, 8, or 10
  • 2.2 GHz or higher processor
  • 4GB RAM  USB 2.0 port or higher
  • Display resolution: 1024 x 768 pixels or higher
    Intel Mac with Parallels/Windows
Available Fluorescence Optics Modules VC-535-403
Excitation / Emission: 470nm/535nm
Example Fluorophores:

  • Acridine Orange
  • CFDA
  • FITC

VC-660-503
Excitation / Emission: 540nm/660nm
Example Fluorophores:

  • Propidium Iodide (PI)
  • Ethidium Bromide
  • 7-AAD

Customer Testimonials

Worth the investment

This instrument [Cellometer K2] is definitely worth the investment. It has a plethora of uses and no...

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Jan 3, 2019

Worth the investment

This instrument [Cellometer K2] is definitely worth the investment. It has a plethora of uses and now serves as the go-to instrument for not only counting cells but for measuring the titer of baculovirus-infected insect cells. Additionally, the customer service representatives are very helpful.
Jan 3, 2019

Consistent and dependable

We use Cellometer K2 for cell counting. This is so convenient and easy to use. Each slide can be use...

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Laboratory of Cellular & Molecular Radiation Oncology, MGH
Dec 27, 2018

Consistent and dependable

We use Cellometer K2 for cell counting. This is so convenient and easy to use. Each slide can be used for two different cell lines and the results show the total number of cells, the number of live cells and dead cells. So we can use this for a lot of experiments which are based on cell numbers and make our results more consistent and dependable.
Laboratory of Cellular & Molecular Radiation Oncology, MGH
Dec 27, 2018

Reproducible results

In comparison to the standard method for manual cell counting, Cellometer K2 helps me in counting ce...

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Dec 11, 2018

Reproducible results

In comparison to the standard method for manual cell counting, Cellometer K2 helps me in counting cells efficiently and quickly with reproducible results. I use it mainly for assessing cell vitality by trypan blue exclusion assay.
Dec 11, 2018

This machine has transformed the way I culture

This machine [Cellometer K2] has transformed the way I culture. No longer do I have to spend hours o...

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Nov 29, 2018

This machine has transformed the way I culture

This machine [Cellometer K2] has transformed the way I culture. No longer do I have to spend hours out of my day to manually could cells. It has changed the way I see cell culture from a menial chore to something I look forward to every day. Cheers NEXCELOM for a job well done!!!!
Nov 29, 2018

Allowed us to enter a new field of cell culture

The K2 has allowed us to enter a new field of cell culture with best pract...

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Nov 21, 2018

Allowed us to enter a new field of cell culture

The K2 has allowed us to enter a new field of cell culture with best practices already in place. Not having to optimize the use of a cell counter will save us a lot of time.
Nov 21, 2018

Very good alternative to flow cytometer

Cellometer K2 is very compact, reliable and easy to use. Our lab uses this counter to count live and...

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Stanford University
Oct 26, 2018

Very good alternative to flow cytometer

Cellometer K2 is very compact, reliable and easy to use. Our lab uses this counter to count live and dead cells for our experiments. It is a very good alternative to flow cytometer for cell cycle and apoptosis studies.
Stanford University
Oct 26, 2018

Easy to use

Cellometer [K2] is easy to use. I like that the program can calculate transfection efficiency, one o...

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Oct 24, 2018

Easy to use

Cellometer [K2] is easy to use. I like that the program can calculate transfection efficiency, one of the things that we do on a normal basis in our lab but we were not able to quantify in the past.
Oct 24, 2018

This has made our life much easier.

We really like this instrument [Cellometer K2]. We were counting cells manually until now. This has ...

more
MD Anderson Cancer Center
Oct 8, 2018

This has made our life much easier.

We really like this instrument [Cellometer K2]. We were counting cells manually until now. This has made our life much easier.
MD Anderson Cancer Center
Oct 8, 2018

Quicker and more accurate

Cell counting is not only quicker and more accurate [with Cellometer K2] but assays that previously ...

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Sep 11, 2018

Quicker and more accurate

Cell counting is not only quicker and more accurate [with Cellometer K2] but assays that previously required a flow cytometer, such as Annexin V/PI apoptosis analysis, can now be completed in our lab!
Sep 11, 2018

Fancy and compact

The design of the instrument is very fancy and compact. It can have its own screen or you can use a ...

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Aug 27, 2018

Fancy and compact

The design of the instrument is very fancy and compact. It can have its own screen or you can use a separate screen. Initially, our lab was using the manual method of cell counting by microscope. But [Cellometer] K2 from Nexcelom has made our life easy and provided reproducible counting.
Aug 27, 2018

Reliable machine

The instrument [Cellometer K2] is being used in our lab for counting different types of immune cells...

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The University of Alabama at Birmingham
Aug 27, 2018

Reliable machine

The instrument [Cellometer K2] is being used in our lab for counting different types of immune cells, mostly neutrophils (but enucleated cells as well). It has been a reliable machine, no problems in general.
The University of Alabama at Birmingham
Aug 27, 2018

Convenience and speed

The convenience and speed of our Nexcelom K2 and Vision Cellometers allowed us to dramatically scale...

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Aug 24, 2018

Convenience and speed

The convenience and speed of our Nexcelom K2 and Vision Cellometers allowed us to dramatically scale up our processing of mononuclear cells. Our sales and technical representatives have always been helpful and responsive to any inquiries.
Aug 24, 2018

Simple and intuitive

[Cellometer] K2 allows us for a faster, reliable and more accurate cell counts that our lab uses so ...

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Northwestern University
Jul 19, 2018

Simple and intuitive

[Cellometer] K2 allows us for a faster, reliable and more accurate cell counts that our lab uses so frequently. The process itself is simple and intuitive compared to the other machine we used to have. Such a great improvement!
Northwestern University
Jul 19, 2018

Customer Publications using Cellometer K2

Our customers include: