RE: [Histonet] Vibratome Considerations
A primary reason to use a Vibratome(TM) is to avoid damage to cell membranes and resulting loss of cellular contents. Do not freeze and rethaw, after using a Vibratome, and you can get more sensitive staining, depending on the what sort of cellular protein you are after (in cytosol, or bound to membranes or other solids) with a Vibratome.
Charles W. Scouten, Ph.D.
5918 Evergreen Blvd.
St. Louis, MO 63134
Ph: 314 522 0300
FAX 314 522 0377
From: firstname.lastname@example.org [mailto:email@example.com] On Behalf Of Geoff McAuliffe
Sent: Tuesday, April 13, 2004 1:03 PM
To: Danielle Zalinski
Cc: HistoNet Listserve
Subject: Re: [Histonet] Vibratome Considerations
I have done immunos on vibratome sections of rat brain. If you use a 12 well tissue culture plate (with its lid) on a shaker table overnight at room temp. you should get good results. You will need to add some Triton or Tween to the primary antibody (and probably subsequent steps) or do something to 'permeabilize' the sections (freeze-thaw or alcohol) so you get good penetration of the antibodies. If you have access to a sliding microtome with a freezing stage you could obtain sections that way.
Danielle Zalinski wrote:
> Hello All,
>Our lab would like to do immunohistochem work in rat brain on vibratome
>sections instead of paraffin. We are able to get sections about
>50-60um. Will incubation with primary antibody overnight be sufficient?
> Is shaking the sections an option? I was hoping some other labs that
>are working in vibratome sections may have some beginners advice.
>Henry Ford Health System
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Geoff McAuliffe, Ph.D.
Neuroscience and Cell Biology
Robert Wood Johnson Medical School
675 Hoes Lane, Piscataway, NJ 08854
voice: (732)-235-4583; fax: -4029
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