MGP Procedure
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From: | rschoonh@sph.unc.edu (by way of histonet) |
To: | histonet@histosearch.com |
Reply-To: | |
Content-Type: | text/plain; charset="iso-8859-1" |
>From one of my "old" files (18+ years ago). I do not have the reference and
the notes are mine. It has been a long time since I did this but I remember
how much fun the I had with the seperation (long story that I don't want to
get into).
METHYL GREEN-PYRONIN STAIN FOR RNA AND
PLASMA CELLS
Procedure:
1. Use Carnoy's-fixed paraffin sections if available. Formalin fixed
tissue is adequate.
2. Run sections down to water.
3. Stain in MGP reagent for 1/2 - 2 min. (longer for old solution)
4. Differentiate in 2 changes of distilled water.
*5. Dehydrate rapidly in 2 changes of acetone, clear in xylol, and
mount in Permount.
*Note: If acetone removes too much of the red stain, slides may be blotted
dry with bibulous paper at this point.
Results: Cytoplasmic and nucleolar ribonucleic acid (RNA): red.
Plasma cell
cytoplasm will also stain red.
Note: Always run a control section fresh fixed liver or
tonsil.
Methyl Green-Pyronin Reagent
Pyronine YÖÖÖÖÖÖ..0.25 gm
Methyl green. . . . . ÖÖÖ.0.75 gm
Phosphate buffer, pH 5.3Ö.100.0 ml
To this add:
0.5% phenol solutionÖÖÖÖ.0.5 ml
1.0% resorcinol (fresh)ÖÖÖ..2.5 ml
Note 1: The Methyl Green will need to be extracted with 3 changes of
chloroform in a separating funnel
Note 2: The MGP reagent is relatively stable. It will take 2-3 days to
ripen and will last 3-6 months if stored at 4 oC.
Phosphate Buffer Mixture
M/5 Disodium phosphate (Na2 HPO4 )ÖÖÖÖÖÖ 52.5 cc
M/10 Citric Acid ÖÖÖÖÖÖÖÖÖÖÖÖÖÖ.. 47.5 cc
Adjust to pH 5.3 (both of the above made up in 25% methyl alcohol to prevent
mold growth).
MGP for Smears
Use control slide.
1. Fix in 10% formalin methanol 10 min.
2. Distilled H2O 2 times.
3 . Air dry 10-15 minutes.
4. MGP reagent 5 minutes.
5.. Distilled H2O 2 times
6. Acetone 2 times.
7. 50/50 Acetone/Xylene
8. Xylene
9. Mount in Permount
best regards,
Bob
Robert Schoonhoven
Laboratory of Molecular Carcinogenesis and Mutagenesis
Dept. of Environmental Sciences and Engineering
University of North Carolina
CB#7400
Chapel Hill, NC 27599
Phone
office 919-966-6343
Lab 919-966-6140
Fax 919-966-6123
Don't go around saying the world owes you a living; the world owes you
nothing; it was here first.
Mark Twain [Samuel Langhornne Clemens] (1835-1910)
-- Begin original message --
> From: Louise Burrell <lburrell@pathbox.wustl.edu>
> Date: Mon, 22 Nov 1999 15:36:36 -0600 (CST)
> Subject: Re: Daily Digest
> To: HistoNet Server <histonet@pathology.swmed.edu>
>
> dear all-please URGENTLY email me with MGP stain for RNA and DNA---thanks
> ahead of time----i need it sooner than yesterday-
>
> With warmest regards,
> Louise "Beezie" Burrell
> Chief Technologist-Cancer Center
> Tissue Procurement Core Facility
> Campus Box #8056
> Washington University School of Medicine
> St. Louis, MO. 63110
> Ph: 314-454-7615
> Fax: 314-454-5525
>
>
>
-- End original message --
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