The best assay for apoptotic cells is to use Anexin V and Propidium iodide (PI) double staining and then running the cells through a flow cytometer. The Anexin V stains the exteriorized phosphatidyl serines (moved from inside to outside the cell membrane), which occurs in the earliest stages of apoptosis. The PI is excluded from healthy cells but stains the DNA if the cell membrane is not intact (i.e., as in necrosis). Hence, your apoptotic cells are Anexin V positive and PI negative. Your healthy cells are negative for both, and your necrosing cells are positive for both. The protocols are available from almost all companies that sell flow cytometers. A sorting flow cytometer will let you collect the healthy cells for further use.

Many thanks Christopher for your kind suggestions.

Is it also possible to visualize the tissues/cells by binocular / Fluorescence Microscopy instead of running the tissues/cells in Flow Cytometry after both staining?

Thank you.

I agree with Christopher that annexin V with PI or another viability die is the best approach to identify apoptotic Vs dead cells. I always used flow citometry to assess, I think that microscopy is not fully suitable to analyze this. Another approach to assess apoptosis is to obtain protein and run caspases, and if there is a cleavage means that there is apoptosis. Anyway, I would go for annexin V, which is more quantitative and faster approach.

PS: remember to obtain the floating cells if you perform an annexin V staining

I agree with Christopher and Enrique Javier Arenas Lahuerta .

From my experience the best assay to be used to distinguish between viable and death cells is to use Anexin V and Propidium iodide (PI).

I highly recommend using Caspase 3 ( westrenblooting).

MTT assay is commonly used to evaluate cell viability.

Good luck