|From:||Kimberly Carter <firstname.lastname@example.org>|
I have done LCM on parafin 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 specitic areas with contamimation 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 block. It is a
tedious and long job, but one that I feel is neccasary.
I hope you are abel to get all of your answers from the NIH web site.
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 best
> way of preparing the sections for this technique, in terms of make of slide,
> 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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