Re: Leishmans Stain

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From:"Histomail\\" <>
To:Katie B <>
Date:Fri, 18 Jun 1999 09:35:42 +1000
Content-Type:text/plain; charset="iso-8859-1"

Dear Katie,
Leishman's is not unlike Wright's stain, both in dye components, action and
results. Both work well on peripheral blood smears and do a reaonable job on
Bone Marrows, although Wright's is slightly better.
Recipe for Leishmans viz. 0.2% in Anhydrous Mehtanol (Analar preferred from
either Merck or EM Scientific). The older recipes, require you to titurate
the dye, this is not necessary. To make one litre put about 200mls of
Methanol in 1 Litre conical flask. While swirling contents, add dye powder
to flask, continue to swirl vigorously so that dye is suspended. Add a
magnetic flea to flask and make up to one litre with remaining Methanol.
Cover top with parafilm and place on mixer for at least 2 hours. You can at
this point filter through Whatmans No. 1 or finer and bottle, I prefer to
turn off mixer and let stand overnight and filter the next morning. The
stain is ready for use but will improve after a few months. Bottle in
airtight bottles, pref. amber glass, and keep your bottle size down to what
you can use in a month so as to avoid having a large airspace and risking
It is highly advisable to use Sorensen's buffer instead of tap or distilled
water as this can vary from site to site. pH 6.8 is recommended for blood
smears and for sections and tissue smears I suggest 6.4 if looking for Mast
The fixation step is the application of the neat stain solution and anything
from 6-10 mins is satisfactory, but smears must be well dried. The actual
dilution is bucket chemistry, is roughly 1:1 and performed/mixed on the
slide, either by tilting, gentle blowing or carefully with a disposeable
pippette. Do not exceed 1:3 dilution or precipitation will be a problem.
Staining time will be between 6-8 mins  and rinsed off with water then
Unopened bottles are stable for about 4-5 years if kept cool and in the
I can recommend the Gurr Leishman Powder and the Gurr buffer tablets are
very convenient with an indefinite shelf life. These can be sourced from
Merck P/L direct. Ph. 1800 335 571 in Australia, BDH in UK or EM Science in
US, and Merck P/L Darmstadt for Europe.
I forgot to mention, if you overstain, then let slides stand in buffer till
differentiated, if severely overstained, add 2-4% Methanol to buffer rinse
and rinse quickly with water to stop further differentiation.
Regards Mike Rentsch (Downunder)
-----Original Message-----
From: Katie B <>
To: Histonet Server <>
Date: Friday, 18 June 1999 7:39
Subject: Leishmans Stain

>I have a visiting scientist here (from Scotland?)who is looking at lung
>lavage cytospins from horses.  She wants to do something called a
>Leishmans stain in order to better identify mast cells and basiophils.
>She stains normally with the Leishmans for 2 min then dilutes it with
>di water and stains for 8 min then rinses completely in water.  It's
>not too different from our Diff Quick, but less pink.  This stain is
>what she is acostomed to looking at, but she doesn't have a recipe.
>Anyone out there in Histoland know what this stuff is?
>As always, thanks for all the knowledge out there!
>Catherine "Katie" Bresee Bennett
>Laboratory for Experimental Pathology
>Department of Veterinary Pathology
>Michigan State University
>Do You Yahoo!?
>Get your free address at

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