HTS / HCS Screening siRNA induced apoptosis assays
Assays for apoptosis can be useful in the study of the effects of drugs and inhibitors (e.g. siRNAs) on cells.
One measure of apoptosis is the surface labelling of cells with Annexin V following membrane inversion and exposure of the phosphotidyl serine residues to the lumen. This is usually visualised with a fluorochrome tag on Annexin V.
HCS platforms commonly use a nuclear dye to localise the cellular "events" to be measured. In HCS it is most convenient to perform the assays on live cells, avoiding extra steps, and to use a dye which is excited in the visible range yet which does not have emission overlap with useful fluorochrome labels such as GFP, FITC, Cy3 and so on.
DRAQ5™ fulfills these criteria and its use in such a HCS-based apoptosis assay for siRNAs was described by Mobius et al. The assay was performed on two different (medium throughput/development and high throughput/screening) platforms demonstrating that a DRAQ5-based assay is readily transportable between them.
The combination of the 488 nm excited Annexin V-FITC and 633 nm excited DRAQ5 leaves significant scope for a third parameter emitting in the "red" part of the spectrum e.g. DsRED, mRFP, Texas Red, TRITC, Cy3.
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