[Cytometry] Shift in florescence during the acquisition

Elena Fingerut fingerute at immunovative.co.il
Wed Dec 23 08:00:52 EST 2009

Dear all, 

I am with cancer patient blood samples preserved for 4 to 7 days in
Cyto-chex tubes, by Streck.

I am trying to analyzed changes in percentage of different cell types and
activation markers on these cells during the clinical protocol. The Ab's
(CD45, CD3, CD$, CD56, CD45RA, CD45RO, MHC-II, MHC-I, CD86) are all from
coulter and acquisition done on FC500 MPL with one laser. The panels include
4 colors. I am working with this combinations of Ab&FACS&Cyto-chex tubes for
a while and it worked fine with a normal blood samples and cancer patient.
Recently I strange phenomena appeared. I have several panels for the same
blood samples and during the acquisition I see a shift in florescence in all
cells populations in sam of the panels, but not in the others. Same time it
happens in all Flu's, same time in one or couple. Same time this double
picture appears from beginning of acquisition.. Basically it looks like a
picture below: 

Form me it looks like in-stability of the FADS but the FACS tech support
says that it is not the machine. He suggest the acquisition test: to acquire
samples as following: QC, water, the problematic sample, water, QC, water,
the problematic sample, water, normal sample, water, QC, water, the
problematic sample, water, QC. All the Act's were fine as well as the normal
samples. The problematic samples looked as above. I repeated the staining of
the same sample with the same Ab's mix according to the same protocol and
acquired at the same setting - the results this time looked perfect. So I
can't really correlate this problem to anything, not blood sample, not Ab
mix and not the protocol. 


Can any one help me with this problem?

I'll appreciate any input..


Thanks to all

And happy holidays.







Elena Fingerut, Ph.D

Research Manager

Immunovative Therapies Ltd.

Mobile phone:+972-54-593-0030


 <http://www.immunovative.co.il/> www.immunovative.co.il


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