Cellometer Auto T4 Cell Counter from Nexcelom

February 07, 2013

Overall

Performance

Ease-of-Optimization

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Stanford University
Structural Biology
Post-Doc
Cell count of human CD4+ cells after column isolation. Viable cells are circled green and non-viable cells are red.

Company:

Nexcelom

Product Name:

Cellometer Auto T4

Catalog Number:

CA-T4-MC

Image

We utilize a number of different tissue culture models including epithelial cells (lung and GI) and suspension cells (leukocytes), both primary cells and cell lines. We measure gene expression changes and normalize our data to cell number, so it is important to have accurate cell counts. In addition, we wanted an instrument that would allow us to exclude non-viable cells or cellular debris. We also needed an instrument that would accurately count a wide variety of cell types.

Experimental Design and Results Summary

Application

Cell counting for subsequent gene expression analysis

Starting Material

Lung cells, GI cells, leukocytes, among others

Protocol Overview

This is a slide based system that requires connection of the counter to a laptop or other PC. If viability counts are desired, cells are treated with Trypan blue (10 µl Trypan blue + 10 µl of cells) and then placed on a slide. Each end of the slide contains a chamber where 20 µl of a cell suspension can be added, allowing two sets of samples to be counted with one slide. The slide is inserted into a port on the front of the reader. The cellometer software is opened on the connected PC, and the cells are brought into focus manually by turning a focus knob on the cell counter as a real-time image is displayed on the computer screen. The type of cell to be counted is selected from a drop down menu (which contains a very large sample of cell types) and counting is initiated by clicking a “count” icon. Counting is relatively fast and depends on the number of cells present. Typically, we observe a 5-10 second time frame.

Tips

If only one chamber of the slide is used, it can be wrapped in a kimwipe and saved for future use. The cell counts do not reflect the dilution performed when staining with Trypan blue, so the user needs to remember to multiply the counts obtained in this case by two.

Results Summary

We contacted the company for a 2-week trial, and they shipped the instrument and a laptop with the instrument software pre-loaded. Over the course of the trial, we counted a variety of cells using Trypan blue to label non-viable cells. We compared cell counts to manual counting under a microscope and flow cytometry when possible. We found that cell counts of CD4+ cells obtained from PBMCs by magnetic bead separation were within 5-10% of counts obtained by flow cytometry using a Guava PCA flow cytometer. The optimal cell count range is 2.5 x 10^5 to 1.0 x 10^7. For most of our counts, we had no problem with a lower limit of 1 x 10^5cells/ml and an upper limit of 5 x 10^7 cells/ml . Cell counts are displayed on the computer screen as total number of cells/ml, with the total number of viable and non-viable cells in different boxes. Out of a few models tested, the Cellometer gave accurate results which were reproducible. It had no problem counting different types of cells, and it has pre-programmed options for different primary or cell lines. There was no difficulty counting irregularly shaped cells.

Features Summary

One of the nice features of the instrument was the ability to count clumps of cells. Everything counted can be displayed on screen with a green circle around viable cells and a red circle around non-viable cells. Another nice feature is the ability to display cell size as a histogram. Images can be stored on the computer for future reference.

Additional Notes

We ran into one experiment where the instrument was counting background, likely cellular debris. We contacted tech support and they sent us an updated software file, where after selecting our cell type from the drop down menu on counting, we then had accurate counts of the cells and excluded background.

Image Gallery

Cell count of human CD4+ cells after column isolation. Viable cells are circled green and non-viable cells are red.

Summary

The Good

Accurate counts, ability to count clumped cells and exclude background.

The Bad

Price. In addition to the instrument, slides can be a little on the pricey side.

The Bottom Line

We are happy with our purchase. The instrument has met our expectations.

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