|From:||Kimberly Carter <firstname.lastname@example.org>|
I have done LCM on paraffin embedded tissue for about 2 years. It is not
something I do often, but I get fairly good results. Since at this time I
do not have access to fresh tissue, I have no first hand knowledge of that
procedure. I cut sections at 8-10u. This is thicker than normal, but
produces more DNA. I use a clean distilled water bath and put the section
on plain slides. I do a progressive H&E stain so not to expose the section
to clarifiers and bluing reagents(this would increase the chance of
specimen falling off). I have also found that you need to do the capturing
either the day you stain or the next day. Since you do not cover slip, the
tissue dries out and more tissue pulls off the slide than what you want. I
work in a research lab at a major university with some of our grants funded
by the National Institute of Health.(NIH) They have a great web site with
all the information you will need on LCM. It is www.nih.gov. You type in
LCM into the search engine on the home page. It is a great resource. A big
thing to remember is that in doing this procedure is that you want DNA and
/or RNA from specific areas without contamination from surrounding areas or
previous blocks. So I clean everything between every block. Change blades,
water bath, tear down microtome to clean all areas and change staining
solution between every block so not to transfer DNA/RNA to next specimen.
It is a tedious and long job, but one that I feel is necessary.
I hope you are abel to get all of your answers from the NIH web site.
OSU Medical Center
Comprehensive Cancer Center
Garry Ashton wrote:
> Dear all,
> Following on from the couple of e-mails about laser capture
> microdisssection, I am interested in what people have found to be the
> way of preparing the sections for this technique, in terms of make of
> section thickness, drying of sections etc.
> In the past we have cut 4um sections, placed on plain unsubbed superfrost
> slides and dried for 1hr at 37 degrees with varying success.
> Any information as ever gratefully received.
> Garry Ashton
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