[Histonet] Suggestion for Dual staining Kit/

Katy Milne kmilne at bccrc.ca
Fri Nov 4 14:35:12 CDT 2016

Hi Nasir, 

I use Biocare reagents (DAB, Warp Red and Ferengi Blue) for my multicolour stuff.

Even though the Vector blue fluoresces you should still see what you need to see in brightfield.  If you're worried about losing your signal through solvents you could probably just airdry before coverslipping, that's what I do (using Ecomount).  The xyelene is only a problem after you put the chromogen on, not before.

Looks like you have 2 rabbit and 1 rat Ab for your Abs, it may be possible to do a triple stain with that with a rat and rabbit in the first round and the other rabbit in the 2nd or you should be able to combine at least a rat and rabbit in a double simultaneous or 2 rabbits in a double sequential stain depending on your Abs and how robust they are.  If you don't have access to spectral imaging though it may be safer going with doubles so you can pull everything apart by eye.

Is that CD8 Ab behaving ok for you?  Looks like it has a ton of background in the images.  If you're having issues with it you may be able to combine your CD4 Ab with the CD3 SP7 clone to show the presumptive CD8s.

I haven't worked with any of those Abs but I do do multicolour IHC in both human and mouse tissues so if you need help sorting out protocols let me know, of course I can only speak to Biocare reagents for this as that's what I usually use.

Katy Milne
Deeley Research Centre

Message: 2
Date: Fri, 4 Nov 2016 15:36:54 +0000 (UTC)
From: Nasir Abbas <shinwari_1 at yahoo.com>
To: "histonet at lists.utsouthwestern.edu"
	<histonet at lists.utsouthwestern.edu>
Subject: [Histonet] Suggestion for Dual staining Kit/reagent
Message-ID: <735822620.281524.1478273814874 at mail.yahoo.com>
Content-Type: text/plain; charset=UTF-8

Hi All
Can some one suggest dual staining reagents for IHC-P (mouse liver tissue) for my primary antibodies: ab183685, GTX53126 and ab25377.I was suggested vectors labs products (following table) but I am not clear about Vector Blue reagent SK-5300 that whether it can be used in IHC-P, because it features shows it is fluorescent. Moreover it is sensitive to xylene and Histoclear may be used as a clearing reagent in this case, which makes me even more worried. Any suggestion and tips for my dual staining?Table:
| Pri-Ab | Vector Lab Products- Suggested |
| ab183685 | MP-7401-15 |
| GTX53126 | SK-4105 |
| ab25377 | MP-5404-15 |
| ? | SK-5300 |

Sincerely?Nasir Abbas

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