[Histonet] glyoxal fixation

[RSRICHMOND <@t> aol.com]

The people at Anatech, makers of Prefer fixative, have published a review of 
glyoxal fixation that every pathologist and histotechnologist ought to read. 
This working surgical pathologist would like to add - and solicit - some 
comments on Histonet. 

"Glyoxal Fixation and Its Relationship to Immunohistochemistry". Richard W. 
Dapson, Ada T. Feldman, and Dee Wolfe. Anatech Limited, Battle Creek MI. The 
Journal of Histotechnology June 2006;29:65-76.

I don't want to change, but I think we all need to be prepared for the day 
when a manager walks into our laboratory, or a letter from a regulatory agency 
arrives in the mail, telling us that we have to get rid of formaldehyde right 
now. Probably glyoxal is the only acceptable substitute, and we all need to 
have a look at it. I have a number of questions.

Interchangeability of glyoxal products: Prefer is described as a buffered 
solution of glyoxal with a pH of about 4. The formula is a trade secret. 
Competing glyoxal products probably also have trade-secret formulas. So can a lab 
change brands of buffered glyoxal without problems, or does it have to stay with a 
particular brand with its trade-secret buffering? - We've seen a similar 
problem with distilling aliphatic xylene substitutes: every one of them requires a 
separate distillation routine, at least on a spinning-band still. - I'll 
leave it to John Kiernan to comment on the appropriateness of trade-secret 
reagents in histopathology.

Limited time in the fixative: Tissue can be left in neutral buffered formalin 
for quite a long time and still be stainable, but tissue stored in glyoxal 
becomes unstainable after about two weeks. Can glyoxal fixed tissue be 
transferred to 70% ethanol for more prolonged storage? - A very occasional surgical 
specimen requires additional blocks after a week - a bigger problem will be the 
pathologist who doesn't trim his autopsies promptly.

Transition period: A laboratory changing to glyoxal would have to keep IHC 
procedures for both fixatives working for some time. There would have to be some 
way to identify whether a block was fixed in formaldehyde or glyoxal.

Eosinophilia: One ought to be able to distinguish eosinophils from 
neutrophils in tissue sections by nuclear morphology, without having to see granules. 
But quantitation of eosinophils - needed in an increasing number of GI biopsy 
situations - could be a problem. We might need an IHC for eosinophils in some of 
these settings.

Lysis of erythrocytes: Not much of a problem, since we're used to it with 
acid fixatives anyway.

Breast cancer: Elimination of nuclear bubble artifact in breast biopsy 
specimens may raise the apparent nuclear grade of tumors, and thus increase 
Nottingham (Elston-Ellis) scores. 

Prostate biopsies: I'd want to see some prostate biopsies - is somebody from 
OURLab in Nashville still on this list? - with formaldehyde fixation, nucleoli 
are a strong criterion of malignancy, and if glyoxal fixation demonstrates 
nucleoli in benign ductal epithelium, this criterion is lost.

Bob Richmond
Knoxville TN and Gastonia NC