[Histonet] HELP! Need some old fashioned histology advice

[Mejia, Mary]

OK Jay, I'm going to have to make this drink of your - thanks for the recipe.
How thick were these whole rabbit mounted sections?  Were the giant glass
slides gelatin subbed?

I was thinking of mounting a couple of these large 100um brain sections & see
how good penetration is for the antibodies. I was also thinking of adding 1% DMSO
in all the washes - don't about the antibodies though.  Any thoughts on the latter?
Any other ideas?

Best
Maria

________________________________
From: Jay Lundgren [jaylundgren <@t> gmail.com]
Sent: Sunday, January 04, 2015 9:58 AM
To: Maria Mejia
Cc: histonet; Mejia, Mary
Subject: Re: [Histonet] HELP! Need some old fashioned histology advice

I can help with the old fashioned advice:


  *   1 scant teaspoon simple syrup
  *   2 dashes Angostura Bitters, plus more to taste
  *   1 half dollar–sized slice orange peel, including pith
  *   2 ounces good-quality rye or bourbon
  *   1 maraschino cherry

     As for the Histology, is there any reason you cannot mount the sections onto glass slides?  When I was working at Genentech they were cutting frozen sections through whole rabbits and mounting the sections on (giant) glass slides.  I think that rolling the tissue up, inserting it, and then removing it from a glass tube would destroy the tissue.

                                      Sincerely,

                                            Jay A. Lundgren, M.S., HTL (ASCP)

On Sat, Jan 3, 2015 at 8:01 PM, Maria Mejia <mbmphoto <@t> gmail.com<mailto:mbmphoto <@t> gmail.com>> wrote:
First, the very best of holidays to everyone.

Now for the histology part.   Our lab's focus is on the early stages of Alzheimer's Disease in the Brainstem
using celloidin processing & embedding for IHC staining.  This year, our lab will be receiving 6 post-mortem
whole human brains (1 every other month).  After fixation, processing & celloidin embedding, the whole brain
will be serially cut at 100um thick.  Each brain section will be 5 inches x 4.5 inches in size.

I will given 250 of these whole brain sections to stain for tau IHC...that's 1500 whole brain sections/year!!!
1) Does anyone have experience doing manual IHC staining of large free-floating brain sections?
2) What type of staining tools, dishes or other essential equipment can anyone recommend?
3) What's the most efficient way to stain 250 sections for batch IHC staining - such as transferring batch
sections (maybe 5-10) from reagent to reagent?
4) What type of batch apparatus to use?

As for the antibody & ABC steps, I was thinking of placing each section inside a large glass cigar tube
(yep, people use large glass tubes with fitted cap to store cigars), with 5ml of antibody or ABC reagent & gently agitate on
a shaker/rotator at room temp during the incubation.  Does anyone have ideas on this?

Please, any ideas, suggestions or recommendation anyone can provide will be most greatly appreciated.

Best regards
Maria Mejia
UCSF
Department of Neurology
San Francisco, CA


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