[Histonet] IHC on paraformaldehyde-fixed

[Andrea Hooper]

Good point. I never tested it directly side-by-side to check for the 
effects of 1% methanol directly. It could be old-fashioned paranoia. 
However, it was a variable I wanted to omit as I know methanol (we 
are talking 100%) has deleterious effects to some mouse antigens. 
Therefore, it wasn't something I wanted to risk ... particularly 
since 95% of what I do now is bone/hematopoietic tissue work anyway 
which can be a nightmare to begin with.




>Interesting point.
>Since 10% buffered formalin (made from the concentrated 38%
>formaldehyde) contain about 1% methanol, has it been shown that this has
>a deleterious effect on ANY antigens or are we expecting this worse case
>senario as being the norm?
>
>I am not aware of any antigens (or antigen-antibody combination) that
>has been badly effected by 10% formalin that is NOT effected by 10%
>formaldehyde. Are you aware of any??
>
>Regards
>
>Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
>Laboratory Manager & Senior Scientist
>Tel: 612 9845 3306
>Fax: 612 9845 3318
>the children's hospital at westmead
>Cnr Hawkesbury Road and Hainsworth Street, Westmead
>Locked Bag 4001, Westmead NSW 2145, AUSTRALIA
>
>
>
>
>-----Original Message-----
>From: anh2006 <@t> med.cornell.edu [mailto:anh2006 <@t> med.cornell.edu]
>Sent: Friday, 5 December 2008 1:31 AM
>To: Tony Henwood; Jan Shivers; histonet
>Subject: Re: [Histonet] IHC on paraformaldehyde-fixed
>
>
>So true. However, be aware that 10% neutral buffered formalin we use has
>methanol in it which may affect certain antigens so there may be some
>difference in staining (hence why for mouse work we now only use 4% PFA
>in pure PBS). It is good to be aware of the other ingredients in your
>fixative solutions, whether commercially prepared or a homemaede recipe,
>as it isn't only the formaldehyde fixative which can make a difference.
>
>
>-----Original Message-----
>From: Tony Henwood <AnthonyH <@t> chw.edu.au>
>
>Date: Thu, 04 Dec 2008 09:35:09
>To: Jan Shivers<shive003 <@t> umn.edu>;
>histonet<histonet <@t> lists.utsouthwestern.edu>
>Subject: RE: [Histonet] IHC on paraformaldehyde-fixed
>
>
>Gee I hate the term paraformaldehyde (as many of you probably know)
>
>This is an example of how confusion of terms can cause unnecessary work.
>Is "4% paraformaldehyde" different from 4 % formaldehyde?
>
>No
>
>Should any procedure done to tissues fixed in "4% paraformaldehyde" give
>results different to those fixed in 4% formaldehyde or 10% formalin?
>
>No since they are the same thing.
>
>
>As Manoonkitiwongsa and Schultz (Histochem J 34: 365-367, 2002) state
>when paraformaldehyde actually becomes a fixative, it is no longer
>paraformaldehyde by chemistry or fixation capacity. Rather, it is
>formaldehyde in water without methanol or any other stabiliser. Without
>heat and an alkaline environment, paraformaldehyde in water is simply a
>paraformaldehyde suspension with little fixation capacity. If the
>fixative is prepared from paraformaldehyde then it should be termed 4%
>formaldehyde freshly prepared from paraformaldehyde. If a concentrated
>formalin solution (40% formaldehyde) is used, then it should be termed
>10% formalin.
>
>If you do a search on Histonet for paraformaldehye, you will find that
>this topic has been extensively discussed.
>
>Regards
>
>Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC) Laboratory
>Manager & Senior Scientist
>Tel: 612 9845 3306
>Fax: 612 9845 3318
>the children's hospital at westmead
>Cnr Hawkesbury Road and Hainsworth Street, Westmead
>Locked Bag 4001, Westmead NSW 2145, AUSTRALIA
>
>
>
>
>-----Original Message-----
>From: histonet-bounces <@t> lists.utsouthwestern.edu
>[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Jan
>Shivers
>Sent: Thursday, 4 December 2008 8:34 AM
>To: histonet
>Subject: [Histonet] IHC on paraformaldehyde-fixed
>
>
>Has anyone ever done IHC on parafomaldehyde-fixed tissues, and if so,
>how well did it work?  Will the same antigen-retrieval methods used with
>formalin-fixed tissue be applicable?
>
>I'm asking for an investigator, who already has his tissues fixed in
>paraformaldehyde.
>
>Jan Shivers
>Senior Scientist
>Pathology Teaching Program
>Histology/IHC/EM Section Head
>University of Minnesota
>Veterinary Diagnostic Laboratory
>1333 Gortner Ave.
>St. Paul, MN  55108
>612-624-7297
shive003 <@t> umn.edu
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