In the meanwhile, I found these:

Lee's Methylene Blue-Basic Fuchsin Stain

Stock Solutions

1. 4 x 10-3 M methylene blue (CI basic blue 9; CI 52015): 0.128 g methylene blue in 100 mL distilled water.

2. 4 x 10-3 M basic fuchsin (CI basic red 9; CI 42500): 0.1296 g basic fuchsin in 100 mL distilled water.

3. 0.2 M phosphate buffer, pH 7.2 to 8.0.

Staining Solutions

1. 4 x 10-3 M methylene blue 12 mL

2. 4 x 10-3 M basic fuchsin 12 mL

3. 0.2M phosphate buffer 21 mL

4. Ethanol (95% or absolute) 15 mL

Staining Procedure

Stain is prepared fresh daily.

1. Mix well.

2. Stain slides for 15 seconds.

3. Wash briefly in running water.

4. Blot dry with bibulous paper.

5. Mount from xylene with synthetic mounting media.

Modified Lee's Stain

This modification of Lee's stain was developed in our laboratory to accentuate nuclear staining.

Stock Solutions

1. 4 x 10-3 M toluidine blue (CI 52040, FW 305.83): 0.1245 g in 100 mL distilled water.

2. 4 x 10 -3 M basic fuchsin (CI 42500, FW 323.815, CI basic red 9): 0.1296 g in 100 mL distilled water.

3. 4 x 10-2 M methylene blue (CI 52015, FW 319.851, CI basic blue 9): 1.28 g in 100 mL distilled water.

4. 0.2M phosphate buffer, pH 7.2 to 8.0

Staining Solution A

1. 4 x 10-3 M toluidine blue 12 mL

2. 4 x 10-3 M basic fuchsin 12 mL

3. 0.2 M phosphate buffer 21 mL

4. Ethanol (95% or absolute) 15 mL

Staining Solution B:

1. 4 x 10-2 M methylene blue 6 mL

2. Distilled water 6 mL

3. 4 x 10-3 M basic fuchsin 12 mL

4. 0.2 M phosphate buffer 21 mL

5. Ethanol (95% or absolute) 15 mL

Staining Procedure

1. Stain in Solution A for 45 seconds.

2. Rinse briefly in running tap water.

3. Blot dry with bibulous paper.

4. Stain in Solution B for 25 seconds.

5. Rinse briefly in running tap water.

6. Blot dry with bibulous paper.

7. Mount from xylene with synthetic mounting media.

What do you think about them? May they be good options to replace the first stain with Polychrome I and II?

Dear Barbara Loi ,

you use (KULZER) TECHNOVIT 7100 resin for embedding your specimen and -after sectioning these 'hydrophilic resin' sections - you will stain these sections.

The sections can be used for histological staining and enzyme detection. A variety of different stainings available at:

https://laboratoryresource.com/?navaction=getitem&id=186 :

>>Technovit H7100/H8100

Technovit H7100/H8100 Datasheet covering staining, sectioning and mounting, Glycol Methacrylate embedding for material samples, soft tissues and immunohistochemistry.

Author: ProSciTech Pty Ltd Last Updated: 2015-07-2 http __ > ://...

Dear Wolfgang H. Muss ,

thank you very much for sharing the information.

My sources are:

1st protocol (Methylene blue/azure II/basic fuchsin): HCMR lab at Crete

2nd and 3rd protocols (Lee's and modified Lee's): Anna C. Siler, Dorothy Thomas, Rolland C. Reynolds, and L. Maximilian Buja. Glycol Methacrylate for Histology: Problems and Solutions. LABORATORY MEDICINE • VOL. 15, NO. 5, MAY 1984, pp. 338-341.

All the best

Dear Barbara, thank you so much for the references which I acknowledgedly I am collecting too into my works collection I have in my files... You might know already that in my former active profession I dealt also with staining of (hydrophobic epoxide/s) sections (cf.my “publication profile“, e.g. AMbF-modified after Humphrey & Pittman 1965 (see 'correction' below) which unfortunately also has MetOH as solvent)... would you mind to inform about the ban of ‚methanol‘ in your lab -honestly asking you about the reason...?

Thank you in advance, best regards, whm

(sent from my iPhone)

NB: EDIT for correction 2020-08-01:

>> Humphrey & Pittman 1965 must read: Humphrey & Pittman 1974

Dear Wolfgang H. Muss ,

I don't know in your Country, but in Italy labs that intend to employ methanol need to ask the authorization to the National Customs Office, and demonstrate they have equipped storage rooms to stock it. In this moment, my boss doesn't want to start the procedures, so I am obliged to find alternatives to methanol.

I also found this (https://www.emsdiasum.com/microscopy/technical/datasheet/14653.aspx):

Technovit® 7100 Routine Staining, Enzyme Histochemistry According to Gerrits

HEMATOXYLIN-EOSIN

Staining process

1. Stain the sections in hematoxylin in accordance with Gill* (filtrate the dye solution) 15 min

* After staining with hematoxylin (1) the plastic matric can be decolorized with 0.5 ml of HCL (36%) in ethanol 70%: briefly submerse and then quickly process in tap water (2).

2. Blue in tap water10 min

3. Rinse in aqua dest.

4. Counterstain sections with Eosin 2-5 min

5. Dehydrate through ethanol 96% and 100%

6. Clarify with xylen and cover in Eukitt

Result

Nucleus blue

Basophilic cytoplasm blue

Acidophilic cytoplasm pink

Muscle tissue pink

Connective tissue pink

Solutions

Hematoxylin in accordance with Gill

Hematoxylin (C.I. 75290) 6g

Sodium iodate 0,6g

Aluminum sulphate 52,8g

Aqua dest. 690 ml

Ethylene glycol 250 ml

Glacial acetic acid 60 ml

Eosin

Eosin Y-(alcoholic) C.I. 453800, 5g

Ethanol 96% 100 ml

Glacial acetic acid 2 drops

Best regards

Dear Barbara, thank you for informing about the problem. I shall com back later on to explain the situation in Austria.

Gerrits-protocol works for sure as many other "histological" stains will work on TECHNOVIT 7100... So it is up to you to choose which staining procedure will fit your task(s)...(:-)) best wishes and regards, WHM.

Barbara Loi Almost certainly, ethanol will work well as an adequate substitute. Indeed I've only ever used methylene blue in EtOH. Indeed, your recipe looks over-complicated at first glance.

Dear Alan F Rawle ,

thanks a lot. Which recipe I listed is complicated according to you? Where can I make things easier?

Barbara Loi The recipe I'm talking about is the one described immediately below your question. See attached. My starting point would be 1% methylene blue in ethanol perhaps let down to 50% in DI water.

You can substitute propanol for methanol in the stain. You will have better staining reactions, although its more expensive but a better substitute

Dear Wolfgang H. Muss is Methylene Blue (MB) cationic? I need to quench green autofluorescence of a slightly hydrophilic, cured photoresist polymer which probably has negative charge residues on its surface. Could you suggest a suitable protocol for MB?

I am using the following product:

https://www.merckmillipore.com/DE/en/product/Methylene-blue-C.I.-52015,MDA_CHEM-159270

Best regards

Muhammad Husnain Ul Abdeen

Please do not hijack this question but open up a new question/discussion. Thank you.

Despite the request of Alan F Rawle to Muhammad Husnain Ul Abdeen to post another - new - question for his answer I would like to

i) answer very briefly to Mohammad (see below) and

ii) would like to ask Mohammad - if he is going to post another , new question regarding the matter - to post in this particular thread the URL of the new post on (proposed title!) 'Is Methylene Blue (MB) cationic and therefore usable to bind to negative charges on surface of cured photoresist polymer?'

Regarding 'Is Methylene Blue (MB) cationic':

First of all:

Admitting that I am not omniscient but had always good books bought by myself during my active work and 'academic' career, in the years of my studies as well as later on used inter-library loan extensively and, since the advent of computer(s) and WWW using the powerful tools of many many literature searches via Google, PubMed, and/or any other Data base(s) I always think it is of value or benefit for others to cite references or sources my 'knowledge' might come....

In this case it is:

'YES, Methylene Blue (MB) IS cationic"

[Quote(s)]:

Chapter 20: THIAZINES / subchapter: Cationic Dyes

„Methylene Blue, C.I. 52015, C.I. Basicblue 9: Is a Diaminothiazine with a small planar conjugated , and of a small overall size. The cation-…- is weakly hydrophilic.(p.299) and, moreover:

Redox indicators (Chapter 8)

„A well understood example is provided by methylene blue, whose cation can be chemically reduced to leucomethylene blue. …. (p-93-)

[End of Quote(s)] -----------------------------------

Retrieved from >> CONN’s 10th Edition quenching+autofluorescence+"photoresist polymer"< or similar combination of search words/phrases and - as a last resort - ask a sophisticated (polymer-) chemist.