Hematology Lessons Best Lab: 7-AAD Viability Testing

7-AAD Viability Testing

Background Information

7-Aminoactinomycin D which is most commonly abbreviated as 7-AAD is a chemical compound that has florescence capability upon intercalation with double-stranded DNA. 7-AAD is manufactured in a manner where it selectively binds to the GC regions of DNA, hence making this nucleic acid stain an effective useful marker in assessing cell viability in multicolor fluorescence microscopy and flow cytometry. 

Principle of 7-AAD Viability Testing
As explained per above, 7-AAD can efficiently separate cells into viable and non-viable. 

Viable cells have cell membranes that are fully intact and are selectively permeable. 7-AAD is a complex compound that does not readily penetrate the cell membranes of viable cells, hence when these viable cells pass through the flow cytometer, there will be no florescence detected and will remain dark.

However, non-viable cells have compromised cell membranes, which will facilitate the penetration of the 7-AAD compounds to intercalate with the intracellular DNA strands. Therefore when these non-viable cells pass through the flow cytometer, the fluorescent capability of the 7-AAD compound will be excited by the laser, resulting in light emission to be detected.

RIGHT DIAGRAM: Dead cells appear brightly lit as shown in the red box as there will be a high concentration of DNA bounded by the 7-AAD compound. Live cells (Viable) appear dimly lit on the other hand

LEFT DIAGRAM: A histogram illustrating the intensity of the brightness of the dead cells (RED BOX) in comparison with the cells that are “viable” 

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