[Histonet] Re: IHC Question

[Johnson, Teri]

Dear Ellen,

You wrote: "I'm interested in performing IHC on a mouse ligament.    Given their minuscule size,  in lieu of paraffin or cryostat embedding and sectioning, we would like to embed in plastic and section at 1 micrometer.   Things we are interested in detecting include collagen type 1 and III, CD31, Arginase 1 and iNOS---to name a few.   Is there a plastic out there that is recommended for Immuno work?   I have performed a brief literature search and have come across Immunobed, JV-4 as well as LR White and Lowicryl.   Is anyone familiar with any of these and whether I would have any success?    I am familiar with Epon-Araldite embedding and sectioning with glass knives, but have never used any of the above plastics.
Thanks for any info"

I think you have a typo, it shoudl be JB-4 and not JV-4. JB-4 is not suitable for IHC.

Any Methyl methacrylate or polymethyl methacrylate formulas should work for you. In your list, Immunobed, LR White, and Lowicryl would all be suitable.

Sectioning and mounting these are different from usual mounting methods. You will need to wet the block face with alcohol for each section. You will put the cut section on a glass slide flooded with alcohol and stretch it, pulling out wrinkles. It will need to be covered with a plastic film, rolled with a roller, and put into a press of some kind and dried overnight. It helps to watch someone else do this first before you try doing it on your own.

We have limited experience with this. Perhaps one of the hard tissue folks will chime in here with some great resource to help you.

Good luck!

Teri Johnson, HT(ASCP)QIHC
Managing Director, Histology Facility
Stowers Institute for Medical Research
Kansas City, MO