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flow cytometry

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#1 waddle



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Posted 15 November 2004 - 08:18 AM

I am trying to identify the different cell populations in spleen from C57 mice.
It is the first time I am using a cytometer and nobody in the lab is able to help me.
I count the white cells with the cytmter after lysis of the red cells and it is OK. Then I use the FCblock, the antibodies, do the washes...
the problem is when I go back to the cytometer, I don't have cells in my gate of interest anymore (less than 20%), and the rest are events with a very high side scatter and a very low forward scatter (this means that all the cells are stuck together).
I use a solution of PBS (free of CaCl2 and MgCl2, from invitrogen) to which SVF and azide was added to reach 2% SVF and 0,16% azide.
Can this be the reason why my cells stick to each other ?
Does anybody can tell me what kind of solution I should use ?
thanks a lot

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