Victoria Baker wrote:

> Hi Everyone!
>
> The controversy over fixation and IHC has reared its'
> ugly little head again.
>
> So rather than bang my head against the wall from
> frustration, I thought I'd put it out and see what
> others are doing.
>
> The tissue is either rat or mouse, the size of the
> tissue is usually 5-6mm in diameter and approximately
> 4mm in depth.  This is a research facility where time
> isn't always critical (except when something is needed
> yesterday).
>
> So if anyone is working with this under similar
> circumstances do you think you could take a minute and
> just answer a couple of questions?
>
> 1.  How long do you fix your tissues for? 10 hours,
> >10 hours, 12 to 24 hours.

Depends on the antigen I am interested in. For some antigens 2 hours is
the max., others overnight is fine.

> 2.  Do you do fixation on or off of your processor?

Don't have a processor.

> 3.  Do you use different fixatives for different
> procedures?

Different fixatives for different antigens. Everything depends on the
antigen.

> 4.  How long is your overall processing time and what
> do you use to hold tissue in after fixation?

Hold the tissue in phosphate buffer or sucrose. Processing time varies
with the antigen in question.

> 5.  This one may cause some controversy.....Does
> anyone cap their blocks after they have finished with
> them for storage.

I cap with melted wax, old school.

> 6.  Does anyone use a xylene substitute and which one
> do you use.

I use xylene which I have an enormous surplus of.  Poor results with the
orange-odor products.

    In case I forgot to mention it, everything depends on the antigen
you are looking for.

Geoff
--
**********************************************
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
mcauliff@umdnj.edu
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