Re: Binding buffer and Annexin V

From: Richard Allman (Richard.Allman@velindre-tr.wales.nhs.uk)
Date: Mon Jul 03 2000 - 04:09:14 EST


There are many recipes available. As long as they have sufficient calcium
most of them should be suitable.

We use the following:
Binding buffer =
10 mM Hepes adjusted to pH 7.4
140 mM NaCl
2.5 mM CaCl2

Method:
1. Wash cells in PBS, centrifuge at 250g (10 min).
2. Resuspend cells in 192 ul of binding buffer.
3. Add 8 ul of Annexin V-FITC and incubate in the dark for 10 min.
4. Resuspend cells in 0.5 ml of binding buffer.
5. Add 10 ul of propidium iodide (10 ug/ml).
6. FACS within 1 hour.



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