Live/Dead Staining of Cells for Flow Cytometry

October 20, 2015

Stanford
Pathology
Graduate Student

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Company:

Life Technologies

Product Name:

LIVE/DEAD® Fixable Aqua Dead Cell Stain Kit

Catalog Number:

L34957

Live cells need to be distinguished from dead cells when running flow cytometry, and although DAPI can be used in most cases for this distinction, DAPI isn't compatible with stainings that require cell permeabilization. This is why we turned to the Live/Dead dyes.

Experimental Design and Results Summary

Application

Flow Cytometry

Starting Material

Single cell suspension of mouse small intestine

Protocol Overview

Mouse small intestine cells were digested via Liberase-digestion to achieve a single-cell suspension. Cells were then incubated with FcBlock, stained with the Live/Dead dye (1:100) and primary antibodies, then run on the flow cytometer.

Tips

Sometimes a higher concentration of the Live/Dead dye may be necessary for much clearer staining (depending on the cell concentration and cell type).

Results Summary

Live and dead cells are separated relatively well with the Live/Dead dye.

Additional Notes

The staining shown here was done in PBS with 2% serum. Staining may be better with the recommended protocol (staining at a higher dilution in just PBS).

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Summary

The Good

Stains allows discrimination of live cells vs dead cells, and is compatible with intracellular staining.

The Bad

Sometimes the separation is not very clear and may require more of the dye for better separation.

The Bottom Line

Overall, the product is great for distinguishing between live and dead cells, especially when intracellular staining is required.

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