Re: Specific Hb Antigen (? h'globin, etc)
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From: | "J. A. Kiernan" <jkiernan@julian.uwo.ca> |
To: | "B.A.Murray" <bamur@alaska.net> |
Reply-To: | |
Date: | Mon, 15 Mar 1999 01:12:15 -0500 (EST) |
Content-Type: | TEXT/PLAIN; charset=US-ASCII |
On Sun, 14 Mar 1999, B.A.Murray wrote:
> One of the clinicians here at the hospital would like for someone to do
> a Specific Hb
> Antigen on some blocks from years ago. He says that if they can be done
> by immuno
> or any other procedure would be fine.
If Hb is haemoglobin, there are two easy ways to stain it
without the trouble and expense of immunohistochemistry.
1. Any acid (anionic) dye at a slightly alkaline pH
(eg. pH 8) will fail to stai most cytoplasmic proteins
and collagen but will still stain Hb strongly. Red
blood cells show very nicely with acid fuchsine done
this way. Remember to wash in slightly acidified water
and go directly to absolute alcohol to minimize
extraction of bound dye. A few other things (Paneth cells,
sperm tails) will also be stained if present.
2, Almost any peroxidase method will detect the peroxidase
activity of haemoglobin. A traditional technique is
the leuco-patent blue method. Diaminobenzidine (DAB) and
hydrogen peroxide, as used in immunohistochemistry, also
reveals the haemoglobin in erythrocytes: often an
annoyance, but for you the desired result. In animal
material there are a few other sources of endogenous
peroxidase (granules in neutrophil leukocytes; also, in
frozen sections of un- or weakly fixed material the
catalase activity of peroxisomes in many cell-types
might be detected. It can cause confusion in some
groups of neurons in the brain.)
In a paraffin or frozen section of something properly fixed in
formaldehyde you should have no difficulty making preparations
in which most or all the stained material is heamoglobin. Cut
the thickest sections you can see through if the object of
this exercise is to examine vasculature.
If Hb is not haemoglobin but ther product of, perhaps, the
_sonic_humbug_ gene (discovered in 1998 and reported in a paper
in Nature by 89 authors, none old enough to know what haemoglobin
is) then the above advice will be u/s. Some HistoNet subscriber
will probably be on your wavelength; they are a wonderfully varied
lot.
John Kiernan
London, Canada.
P.S. Linda Margraf: Just how many histonetters are there now?
You put out a number in the 800s more than a year ago.
This PS will find out if you read every message right
down to the last line. I hope you don't!
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