Rick While not wanting to claim to be a guru, I have done some fixation experiments with annexin V. I have found that supplementing FACSlyse with Ca++ to 1 mM is required to keep the annexin signal, I assume this is true for any formaldehyde based fixative. In my experience the fixation does not actually fix the annexin and all washes also need to be done in divalent cation containing buffers (I used HBSS). The loss of signal has been quite manageable under these conditions. This is my personal experience, not advice from BDIS. Glad to hear any different observations. Tom Frey _______________________________________________________________________________ Subject: Annexin V and fixation From: RJasman@ICOS.com at INTERNET Date: 12/17/97 11:18 AM For you gurus of apoptosis: I have a client that uses Annexin V to monitor apoptosis in mouse cell lines currently. He has expressed an interest in moving to human cells with his experiments. Our current SOP for human cells is that they must be fixed to analyze by flow. He has tried fixation after Annexin V staining in the past and found that the fixation destroyed the signal (binding?) of the annexin. Has anyone been successful in fixing cells after annexin V staining? Do you have a method? Thanks, Rick Jasman ICOS Corporation
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