I had a hard time returning this reply to the originator.
---------------------- Forwarded by Rande Kline/EMI/Merck on 05/25/99 08:49
AM ---------------------------


Rande Kline
05/18/99 11:08 AM

To:   "NRWillis" <NRWillis@dial.pipex.com>
cc:   "HistoNet Server" <HistoNet@Pathology.swmed.edu> AT
      INTERNET-MAIL@ccMail
Subject:  RE: More on Giemsa  (Document link not converted)

We get a lot of questions involving Wright/Wright-Giemsa/Giemsa staining of
peripheral bloods.  I talk to a lot of hematology labs.

Fixation in methanol for peripheral blood smears should only require no
more than one minute. We recommend 30 seconds.  It may be a good idea in
high humidity areas to use two changes of methanol. Discard the first
change and rotate the second change into the first position as needed.
This would be dependent on how many smears are being stained and once
again, the humidity.

A dilution such as described below is only stable for about 4 hours.  If
using coplin jars, try to keep the lids on during staining.

I get mixed comments on usage.  Most use a combination of stock stain and
stain:buffer solution using pH buffers from 6.4 to 7.1.  Some like the
stain better after the stock stain solution is a day old, some change the
stain daily.  Like the H&E  stain, this stain is very subjective.

I just have to do this plug.  EM Science has a large line of hematology
stains. The one I like to recommend for histology is a Giemsa Solution
imported from Germany.  It works great on paraffin, bone marrow aspirates,
and peripheral blood smears. The catalog # is 9204/4.  This stain solution
is in the Boon and Kok, Microwave Cookbook of Pathology.  Unfortunately, it
is not due in until June.  If anyone is interested and needs the name of a
distributor call me at 800-222-0342x443 or e-mail me.

Regards,
Rande Kline HT (ASCP)
Technical Services
EM Science












From: "NRWillis" <NRWillis@dial.pipex.com> AT INTERNET-MAIL on 05/17/99
      07:54 PM

To:   "HistoNet Server" <HistoNet@Pathology.swmed.edu> AT
      INTERNET-MAIL@ccMail
cc:    (bcc: Rande Kline/EMI/Merck)
Subject:  RE: Gimesa




Andrea,

        Bryan's formulation works well, but I think fixation in fresh
(emphasis
on
fresh) methanol should be at least 10 minutes.  This is followed by
staining in a 1:10 solution of stock Giemsa:Sorensen's phosphate buffer
(pH6.4 - pH6.8) for 20 minutes.  Rinse in fresh buffer, then two changes 2
minutes each of buffer.  Better nuclear staining can be achieved by
following fixation with a 50:50 mix of May-Grunwald:Sorensen phosphate
buffer.

Diluted Giemsa (and May-Grunwald if used) decay rapidly with time.  They
should be prepared fresh (ideally) before use.  In practice twice daily is
sufficient, but where critical use fresh.

Ingress of atmospheric water into fixative methanol, stock stain solutions
leads ultimately to poor results.

In the US I understand that Wright-Giemsa staining is preferred, again
giving better nuclear and cytoplasmic definition than either stain alone.

Hope this helps

Nigel