Detection of Membrane Changes Using YOPRO1

1. Harvest 1 x 106 cells and wash once in PBS.

2. Resuspend cells in 1 mL of medium with 1 |J,MYO-PRO-1.

3. Incubate cells at room temperature for 20 min.

4. Wash once in PBS and analyze immediately (keep cells on ice until analysis).

5. YO-PRO-1 fluorescence, after 488-nm excitation, is measured between 515 and 545 nm.

Addition of a dead cell discriminator such as PI will enable live cells (YO-PRO-1- and PI-negative) to be separated from apoptotic (YO-PRO-1-positive, PI-negative) and dead (YO-PRO-1-positive, PI-positive) cells. In addition, as cells die and become progressively necrotic, they follow a parabolic course on a dot plot, being initially bright for both dyes and then progressively less so.

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