Calcium orange on FACSCalibur

S Newton S.Newton at sheffield.ac.uk
Tue Jan 22 04:09:10 EST 2008


Hello all

I have attached a protocol that a user is performing for calcium flux. We are
using the FACSCalibur and are using Calcium orange as the cells are also GFP
positive. 

There is no calcium flux in the positive control cell line (which is GFP
negative) and I was wondering if there is anything glaringly obviously
incorrect in the protocol. The user is going to use some other dye (Possibly
Flou 3) and follow another users protocol to check that her cells are ok,
however I am concerned that the 488nm laser is too low a wavelength to
excite the calcium orange (although there is a two log decade shift between
unstained and calcium orange loaded cells in FL2).

http://probes.invitrogen.com/servlets/spectraviewer?fileid1=3013ca

Any advice......

Thanks
Sue

-- 
Sue Newton
Flow Cytometry Technician
DU27. D Floor
Medical School
University of Sheffield
Beech Hill Road
Sheffield
S10 2RX

Phone number: 0114 271 3023
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