Giemsa-11 species-spef. staining

Helene Klapper hk-miami at ix.netcom.com
Fri Jul 7 17:52:30 EST 1995


In <3tjc8p$m02 at osiris.wu-wien.ac.at> bunka at i112syw1.vu-wien.ac.at
(Sebastian Bunka) writes: 
>
>
>Hi,
>anyone on the net that has done the Giemsa-11 technique for species
specific
>chromosome differentiation ?
>
>We tried this for differentiation of Mouse-Cat heterohybridomas, but
the
>stain does'nt work for us. Any ideas or comments ?
>
>From the original paper:
>C. Buys, G. Aanstoot, and A. Nienhaus, 1984, Histochem. 81:465-468:
>
>"Oxidizing Methylene blue and adding the reaction products to Eosin Y
and
>Azure B makes possible a highly reliable Giemsa-11 technique for
>discrimination of chromosomes in hybrid cells according to their
parental
>origin..."
>
>Thanks in advance for any comments and tips!!
>
>Cheers, Sebastian
>--
>
>;-)
>----------------------------------------------------------------------
-
>Sebastian Bunka			ph. (+43-1) 711 55 301
>Institute of			FAX (+43-1) 713 68 95
>Medical Chemistry		email: Sebastian.Bunka at vu-wien.ac.at
>University of Veterinary Medicine - Vienna - Austria
>----------------------------------------------------------------------
-
>
I have several protocols for this, each somewhat different than the
others, but I have gotten none of them to work.  A friend of mine who
is an expert in such matters tells me that the pH is CRITICAL - that it
has to be exact, and that it is temperature dependent.  Most of my
protocols involve doing it in a waterbath (above room temp - I don't
remember if it's 37 degrees or what...my notes are in the lab and I am
at home) so she has told me that I need to take the pH of the soln
while it is in the waterbath.  If you take it out to pH it, the temp
changes and therefore, the pH is no longer accurate.  I hope this helps
somewhat.  Helene



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