MGP Procedure

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>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


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

	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,
Robert Schoonhoven
Laboratory of Molecular Carcinogenesis and Mutagenesis
Dept. of Environmental Sciences and Engineering
University of North Carolina
Chapel Hill, NC 27599
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 <>
> Date: Mon, 22 Nov 1999 15:36:36 -0600 (CST)
> Subject: Re: Daily Digest
> To: HistoNet Server <>
> 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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