Hello Johana...
Annexin V labeling can be readily combined with the PhiPhiLux caspase
assays from Oncoimmunin, since both assays are carried out in
"viable" (i.e. unfixed cells). The method is
described in...
__________________
Telford, W.G., Komoriya, A. and Packard, B.Z. (2004)
Analysis of apoptosis by flow and image cytometry. In
Methods in Molecular Biology Volume 263, Flow Cytometry Protocols, 2nd
Edition, T.S. Hawley and R.G. Hawley, eds., Humana Press, London, UK.,
pp. 141-159. (I can send you a copy of this book chapter, which is
a little hard to find).
Telford, W.G., Komoriya, A. and Packard, B.Z. (2002). Detection of
localized caspase activity in early apoptotic cells by laser scanning
cytometry. Cytometry 47, 81-88.
__________________
The method is quite simple - you label the cells with the caspase
substrate, and add the annexin V for the last 15 minutes of the substrate
incubation. The cells are then washed once and a membrane
permeability dye (i.e. PI) is added prior to analysis. It gives a
nice multidimensional picture of the apoptotic process.
The methodology is also described on our website at
http://home.ncifcrf.gov/ccr/flo
wcore/index.
htm
under the Projects section (direct link is
http://home.ncifcrf.gov/ccr/flow
core/caspase3.htm). Oncoimmunin's web site is at
http://www.phiphilux.com .
Enjoy,
Bill
At 02:23 PM 7/28/2004 -0400, Melendez, Johana wrote:
Helo Flowers:
I will like to know if anybody out there can send me a protocol or a
reference for Annexin V staining combined with caspase 3. I
found references of people discussing results , but I have not found a protocol
yet. Your help will be greatly appreciated.
Johana
Johana Melendez, MS
Research Associate
Flow Cytometry Laboratory
H. Lee Moffitt Cancer Center
12902 Magnolia Dr.
Tampa, Florida 33612-9497
Phone (813) 745-6610
Lab. phone (813) 972-8400 x. 2005
Fax (813) 903-7140
<<Melendez, Johana.vcf>>
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Received on Tue Aug 3 14:18:00 2004
