Safranin working solution?

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Raul
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Safranin working solution?

#1 Post by Raul » Mon Jan 25, 2016 10:13 am

Hi guys, I recently bought Safranin as a powder, and I would like some advice on how to make the solution (if aqueous or alcoholic) what proportions should it have and what concentration of the dye. I am looking to do Gram stain but also general nuclear stains and cartilage stains.

Please if you can tell me a recipe that you tried or know to be working ( because I found some on the internet but all where different)

Regards,
Raul

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Raul
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Re: Safranin working solution?

#2 Post by Raul » Mon Jan 25, 2016 10:14 am

Also if anyone can explain me what are the advantages and disadvantages of working with an aqueous solution or alcoholic solution please.

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75RR
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Re: Safranin working solution?

#3 Post by 75RR » Mon Jan 25, 2016 11:26 am

Can't help directly but you may find something by WALTER DIONI at microscopy-uk.org.uk will help.
http://www.mic-uk.info/cgi-bin/search.c ... m_cat[]=-1
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Peter
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Re: Safranin working solution?

#4 Post by Peter » Mon Jan 25, 2016 7:02 pm

I Raul,
As a counter stain to grams stain safranin is used as a simple 1% aqueous solution.
Hope this helps.
Peter.

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Raul
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Re: Safranin working solution?

#5 Post by Raul » Mon Jan 25, 2016 8:57 pm

Thank you 75RR for the link and thank you Peter for telling me the concentration and type of solution.

Best regards my friends.

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mrsonchus
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Re: Safranin working solution?

#6 Post by mrsonchus » Thu Jan 28, 2016 12:34 am

Hi Raul, Safranin, I use aqueous. 1% is my std mix and in this I stain for between about 3 mins up to 24 hours, depending on how hard I intend to hit the stain with a de-colourizer to differentiate back down to the tissue-parts I would like Safranin to stain only.
Also it may be used at 0.05% if you've a delicate and very selective staining-requirement with which to use it - but this is rare for me to use - I find the 1% very useful and consistent.

The idea with Gram, used as I use it not for the Gram +/- distinction of bacteria, but as a good 2-stain combination for Botanical tissue, is as follows;
'Crystal' ('Gentian') violet is used first but is extremely easy to wash completely out of tissue without the use of 'Lugol's Iodine' to 'fix the stain into the tissues' by combining with it, forming a large molecule that will resist washing from tissues and through membranes (such as plasma-membrane or cell-wall/s) facilitated by this increased size.
Once the Cv is safely into the tissue you're ready for the Safranin, which as a 'regressive' stain is added to overstain then reduced back to a level that you're happy with, that of course allows the Cv to be seen as well in the tissue. Incidentally the Safranin is in my opinion a fine nuclear stain - it can give a less-opaque and therefore more detailed nuclear image I find than say Methylene blue.
Without the use of the Lugol's iodine, you will be highly likely to lose the crystal violet.

This is the simple protocol that I use to good effect on all sorts of tissue (botanical);
ws_my_gram_stain.jpg
ws_my_gram_stain.jpg (86.16 KiB) Viewed 1867 times
Hope this may help a little, although it's really not used for true bacterial Gram-staining by me.. :)
John B

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Raul
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Re: Safranin working solution?

#7 Post by Raul » Fri Jan 29, 2016 12:24 pm

Thank you mrsonchus, that is some really good information (especially from the fact that is experimented by you). I intend to do some Gram stain for bacteria, but mainly I will use Safranin histologically, being very similar with what you described.

Thank you very much to all the people that responded.

Regards,
Raul

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