[Histonet] Reg: Beta-2 minroglobin IHC

[Robin Dean]

Hi all,

 

We are trying to use beta-2 microglobulin IHC staining to ID human cells in
goat joints that were engrafted with human cells some time ago and having
nothing but trouble. Tissue was decalcified in immunodecal, formalin-fixed
paraffin-embedded tissue. Beta-2 MG antibody is a polyclonal rabbit X human
beta-2 antibody from Acris, and we are using DAKO Envision-HRP -con.
secondary with DAB as chromogen.

IgG isotype controls and DAB alone controls look clean.

.         Goat joints are falling off the charged slides with heat-induced
citrate buffer antigen retrieval so went to enzymatic retrieval to keep them
on the slide.

o   Does enzymatic antigen retrieval damage this epitope?? (previous
successful uses of this antibody were with citrate antigen retrieval)

o   Any suggestions on how to keep sections on slides? Sections are kind of
large, but pathologist wants that size.

.         Didn't get hardly any staining at 2.5 ug/ml of the beta-2MG
antibody, but at 5 ug/ml everything stains including all of goat joints. 

.         Are there other or better stains that are used for identifying
human cells engrafted into other species (label uniformly most human cells)?
I saw mention of some mitochondrial antgen antibodies but no specifics.

.         Does Beta-2  antibody label only human cells? Seems like there are
homologous markers in other species that may label. 

Currently we aren't using any blocks except peroxidase block. Thought we
might try a CAS block to see if that will prevent sticking? To non-human
tissue.

 

Any suggestions or help with these problems will be greatly appreciated. We
have to get the stain to work and are running out of ideas.

 

Robin 

 

Robin R. Dean, Ph.D.

Senior Scientist & Study Director

Comparative Biosciences, Inc.

786 Lucerne Dr.

Sunnyvale, CA

(408) 738-8060

robin_dean <@t> compbio.com