Vybrant CFDA SE Cell Tracer Kit Clearly Labels Living Tumor Cells

December 28, 2016

Duke University
Immunology
Predoctoral

Overall

Quality of Results

Ease-of-Optimization

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Company:

Thermo Fisher Molecular Probes

Product Name:

Vybrant CFDA SE Cell Tracer Kit

Catalog Number:

V12883

The goal of this research was to label tumor cells in vitro for use in downstream phagocytosis assays identify to then be analyzed by flow cytometry.

Experimental Design and Results Summary

Application

Flow Cytometry

Starting Material

Primary mouse B cell lymphomas

Protocol Overview

Tumor cells in suspension were collected and then washed. Cells were resuspended in either warm PBS alone or warm PBS containing 100 nM, 500 nM, 1 uM, or 5 uM CFSE prepared according to manufacturer's instructions. Cells were incubate for 15 minutes at 37 degrees Celsius. Cells were then washed and resuspended in warm media and were then incubate for 30 minutes at 37 degrees Celsius to ensure probe modification. After washing tumor cells labelled with CFSE, cells were washed and resuspendd in PBS containing Fc block (diluted 1:250) and LIVE/DEAD (diluted 1:5,000), using 100 uL for one million cells). Cells were incubated on ice in the dark for 20 minutes.

Tips

This reagent will need to be optimized for the particular cell population and flow cytometer being used, because it easily gets too bright for analysis and at high concentrations can kill your cells if the cells need to be incubated longer than just to stain.

Results Summary

Tumor cells were clearly labelled at increasing rates using CFSE. Cells that were unlabelled (empty histogram) were somewhat distinguishable from those incubated with 100 nM CFSE, while cells labelled with increasing concentrations of CFSE were more obviously distinguished (increased shading indicates increasing concentrations, 100 nM, 500 nM, 1 uM, 5 uM).

DOI or PMID #

N/A

Additional Notes

N/A

Image Gallery

Summary

The Good

Cells were brightly labelled by CFSE.

The Bad

It takes longer than your standard stain, and there is some optimization to be worked out based on cell size and viability.

The Bottom Line

Great way to label cells for downstream flow cytometry assays, if there is time and reagents available to optimize.

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