Ghost Dye Violet 510 Live/Dead Analysis

June 02, 2017

Columbia University
Microbiology & Immunology
MD/PhD Student

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Company:

Tonbo Biosciences

Product Name:

Ghost Dye™ Violet 510

Catalog Number:

13-0870

Human PBMCs were used for analyses of memory T cell subsets. The Ghost Dye was employed to differentiate between live and dead cells from the freeze/thaw and liquid nitrogen storage process. The violet 510 color was selected to allow for multiple channel acquisition off the violet laser.

Experimental Design and Results Summary

Application

Flow cytometry

Starting Material

Human PBMCs

Protocol Overview

Human PBMCs were thawed from liquid nitrogen storage and washed with complete media (e.g. 10% FBS in RPMI) before staining for flow cytometry analysis. Cells were then washed once in dPBS and resuspended in Ghost Dye diluted 1:400 or 1:800 in dPBS and incubated for 20 minutes on ice. Cells were then washed once with excess dPBS.

Tips

Stain with fixable amine reactive Ghost Dyes in dPBS separate from antibody staining

Results Summary

Labeled PBMCs were analyzed on a BD Fortessa. Cells in the lymphocyte gate were separated into live and dead gates based on Ghost Dye 510 fluorescence (left panel 1:400 dilution; right panel 1:800). Efficient labeling of the dead cellular fraction was observed with both dilutions of dye tested.

DOI or PMID #

N/A

Additional Notes

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Summary

The Good

Inexpensive, efficient labeling of dead cells; useful fluorescent channel options

The Bad

Significant spillover from the Pacblue/BV421 channel when multi-plexing.

The Bottom Line

The Ghost Dye 510 is an inexpensive flow cytometry reagent that allows for efficient labeling of dead cells and subsequent multi-color analysis. After appropriate compensation, the 510 dye is a particularly useful color option for complex panels requiring the use of multiple colors off the violet laser. Ghost Dye 510 is a reasonable alternative to Thermo Fisher's Live/Dead Aqua reagent.

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