[Histonet] Cytology on live culture cells

From:Aprill Watanabe

Hello Histonetters,

I need some help, I work in a research lab and a colleague is trying to
differentiate live cells.  I need a stain that we can do on live tissue
culture cells to differentiate tumor cells from normal cells.  These are
cell lines being started from primary tumors.  They know the population is
mixed but needs a way to tell how pure the population is.  They read an
article about using DiffEQ or the Pap stain.  I just donšt know how to do
this in a flask or petri dish.  I also am at a loss for which stain would
stain which cell in those 2 stains.

My other problem is that I have another colleague trying to stain live
culture cells with cytokeratin AE1/AE3 from DAKO.  We use MACH 3 mouse
polymer kit from Biocare for the detection.  She has tried it a couple of
times and got no staining at all.  She has tested the polymer kit and it
works.  She is using chamber slides to do the assay.  She fixes with 4% PFA
and does a tritonX digest.  She has played with the concentration of the
tritonX and still got nothing.  She is setting up a few test experiments to
test different fixatives and then to add some sort of retrieval step.  The
problem is we arenšt set up for anything other that autostainer retrieval or
by pressure cooker, and wešre using chamber slides so I canšt put them on
the autostainer.

Any help/suggestions for either problem would be great.  Thanks!

Aprill Watanabe, B.S.
Research Associate
Integrated Cancer Genomics Division
Tissue Microarray Center (TMA)
Translational Genomics Research Institute (TGen)
main: 602-343-8822
Fax: 602-343-8840

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