ethanol treatment vs. plasmid loss

Luis Diaz cascas at umich.edu
Sun Nov 6 17:35:39 EST 1994


I have an mAb that recognizes an intracellular epitope of a T cell
membrane glycoprotein.  I want to clone the protein this mAb recognizes
by transfecting a T cell cDNA library into NIH 3T3 cells and screening by
flow cytommetry (FACS) with our FITC-labeled mAb.  The problem I am
concerned about is that to screen with my mAb we have to first
permeabilize the transfected cells with 70% ethanol.  I am worried that a
successfully transfected plasmid would be lost after ethanol treatment. 
Am I wrong to assume the loss of a plasmid after 70% ethanol treatment or
is this something that is likely to occur.

Luis A. Diaz
Medical Student 
University of Michigan



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