I am trying to dual stain with Hoechst33342 and 7-AAD. Does anyone have any experience using these dyes either individually or together?
I set up an experiment following the nature protocols paper (link below) and did not detect any staining (even though I collected my media over 3 days and stored in the fridge, so I definitely had 'dead' cells).
The only parameters I changed was the amount of dyes added due to the recommendations on the datasheets which came with the dyes I purchased. I used 5ug Hoescht and 0.25ug 7-AAD per test instead of the 1ug Hoechst and 1ug 7-AAD suggested.
I could stain Hoechst for 20-30 min and then 7-AAD for 10 min all on ice also. Does any one have any other protocols they would recommend trialing or any other advice?