[Histonet] Problems with MSB stain

[John Kiernan]

The unduly blue sections are all from Animals 12, 13 and 14. Anything different about these specimens? The blue and red sections surely must look different under the microscope.
 
John Kiernan
Anatomy, UWO
London, Canada
= = =
----- Original Message -----
From: Stephen.Eyres <@t> sanofi-aventis.com
Date: Wednesday, January 13, 2010 10:12
Subject: RE: [Histonet] Problems with MSB stain
To: jkiernan <@t> uwo.ca


> Hi John,
>  
> Many thanks for the rapid response. The slides look fine down the microscope except for a few that macroscopically look blue. However because the MSBs will be sent away for evaluation, we want to send a consistent set of slides. I attach a picture to show the problem. I can also scan a slide if you prefer, but the image size is large. The paper was very helpful.
>  
> Cheers
>  
> Steve
> 



> From: John Kiernan [mailto:jkiernan <@t> uwo.ca] 
> Sent: Wednesday, January 13, 2010 6:23 AM
> To: Eyres, Stephen R&D/GB
> Cc: histonet <@t> lists.utsouthwestern.edu
> Subject: Re: [Histonet] Problems with MSB stain
> 

> What do the slides look like under the microscope? In a normal liver there shouldn't be much collagen (blue) and there should be plenty of cytoplasm (red). If the preceding nuclear stain was too strong, that could put unwanted bluish shades in the cytoplasm. 
>  
> Trichrome methods aren't at their best after formaldehyde fixation; the colours can be improved by soaking the hydrated sections in saturated aqueous picric acid for 30 min or so at 55-60C before staining. Bouin's solution is also used for this purpose, and it is said that the same effect can be achieved with citrate buffer pH4 or with 1% iodine in 2% aqueous KI.  See Yu Y, Chapman CM (2003) Masson trichrome stain: postfixation substitutes. J. Histotechnol. 26: 131-134. I've not yet tried these alternatives to picric acid; has anyone else?
>  
> Lendrum's 1962 paper, which includes colour photos, includes many technical tips. More than 40 years ago a med lab technician in Birmingham, UK, suggested this method to me as a way to impart different colours to gliosis (red) and collagenous scarring (blue) in sections of the brains of frogs and other animals in which I'd made surgical lesions dorsal to the optic chiasma. The MSB method showed these changes very well. I recall that we had to buy about 500 grams of brilliant crystal scarlet 6R because it was then cheaper as an an article of commerce than as a bioogical stain.
>  
> I have appended a PDF of Lendrum et al 1962.  This won't  gwt through to all Histonetters, but it might make it to Stephen.Eyres <@t> sanofi-aventis.com. 
>  
>  
> John Kiernan
> Anatomy, UWO
> London, Canada
> = = =
> ----- Original Message -----
> From: Stephen.Eyres <@t> sanofi-aventis.com
> Date: Tuesday, January 12, 2010 11:06
> Subject: [Histonet] Problems with MSB stain
> To: histonet <@t> lists.utsouthwestern.edu
> 
> > Hi,
> >  
> > I have just encountered a problem I have never seen before and would
> > welcome opinions as to the cause. I work in pharma R&D and had a 
> > requestto perform MSBs on some liver slides. The results varied 
> > within the run,
> > with a distinct difference in the overall colour of the  
> > slides; some
> > appearing blue and others red. The person performing the 
> > staining is
> > experienced, and our neqas scores are good. When we investigated the
> > problem the only difference we could identify was that the bluer 
> > slideswere stained after  40 minutes drying after being 
> > cut, whereas the
> > redder slides were cut the day before. This suggests that maybe slides
> > were incompletely dried and that this affected the staining. Comments
> > please.
> >  
> > Cheers
> >  
> > Steve 
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