A convenient alternative to CFSE staining

January 27, 2015

Dana Farber Cancer Institute
Cancer Immunology & AIDS
Research Fellow

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Company:

eBioscience

Product Name:

Cell Proliferation Dye eFluor® 670

Catalog Number:

65-0840-90

I wanted to measure T cell proliferation, but my cells inherently expressed green fluorescent protein (GFP), which has the same emission spectra as the well-known proliferation dye: CFSE. So, I decided to give this efluor670 proliferation dye a shot. This would be read on the APC channel and thus, would not interfere with GFP emission spectra. The staining procedure is very similar except that you stain at a higher concentration (5uM). The dye works perfectly fine as you can see in the attached figure. Very beautiful peaks of dividing cells are seen in the form of dilution of the dye compared to the negative control which does not show any dilution and thus a single peak. Highly recommended product.

Experimental Design and Results Summary

Application

Flow cytometry

Starting Material

Mouse T cells

Protocol Overview

Single cell suspensions were prepared in PBS and stained with 5uM of this dye at 37 degrees celcius (I used a water bath) for 15 minutes. The staining was immediately stopped by adding media with 10%FBS to the cell suspension. The cells were washed and cultured for 48 hours before analysis on a flow cytometer.

Tips

Make a fresh working stock every time.

Results Summary

The cell proliferation can be visualized as dilution of the dye on a flow cytometer. Each peak represents a population of cells divided a certain number of times (the more the dilution of each peak, the more cell divisions have occurred).

Additional Notes

None

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Summary

The Good

Good price and bright staining; less toxic than CFSE

The Bad

The cells have to be free of any carrier protein, otherwise the dye will not be specific

The Bottom Line

It is a great alternative for measuring cell proliferation even if your 488 channel is occupied.

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