FW: [Histonet] in situ question

[Tony Henwood]

Ruth,

But Paraformaldehyde is the powder.
When you chemically treat it (sodium Hydroxide & heat) it dissolves and
forms formaldehyde. So the correct term is 4% buffered formaldehyde or
if made from concentrated formalin (the liquid form) it is termed
10%buffered formalin (c. 4% buffered formaldehyde).

Paraformaldehyde is poorly soluble in pH neutral water.

It's the terminology that many of us often have issue with, apart from
problems of short fixation and why bother to use paraformaldehyde in the
first place when formalin is safer and does an equivalent if not better
job!

Please refer to the article:

Manoonkitiwongsa & Schultz (2002) Histochem J 34: 365-367

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 




-----Original Message-----
From: Yaskovich, Ruth A (NIH/NIDCR) [E]
[mailto:ryaskovich <@t> dir.nidcr.nih.gov] 
Sent: Thursday, 21 August 2008 12:15 AM
To: Tony Henwood
Subject: RE: [Histonet] in situ question


Tony,
 It's actually a 4% BUFFERED Parformaldehyde. It works a lot better with
a buffer. Ruth N.I.H.

-----Original Message-----
From: Tony Henwood [mailto:AnthonyH <@t> chw.edu.au] 
Sent: Tuesday, August 19, 2008 8:02 PM
To: N Fournier; histonet <@t> lists.utsouthwestern.edu
Subject: RE: [Histonet] in situ question

Some questions,

What is "PARA"?

Is it an something you add to the formaldehyde solution?

Please don't use the term 4% paraformaldehyde, use 4% formaldehyde,
unless you are rolling the tissue in a powder consisting of 4%
paraformaldehyde and 96% talcum powder or some other solid!

Please refer to: Manoonkitiwongsa & Schultz (2002) Histochem J 34:
365-367



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 




-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu
[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of N
Fournier
Sent: Wednesday, 20 August 2008 3:37 AM
To: histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] in situ question


Hi,

I have a few questions regarding in situ hybridization. The protocol
that I am following indicates that they perfused rats with PBS followed
by 4% paraformaledhyde (in PBS). However, I always thought that one
generally extracts the rat brain fresh, flash freeze, section, and then
post-fix in 4% paraformaldehyde before commencing with the in situ
hybridization. I suppose it might not matter since in the end the tissue
is post-fixed in PARA; however, I would like to hear what others with
experience doing in situ think about this procedure. Additionally, does
one always use DEPC treated solutions even for these perfusion steps
(e.g., PBS perfusion, paraformaldehyde perfusion)? Lastly, does one have
to section on a cryostat? Could I simply use a vibrating microtome to
section the tissue instead. 

Thanks for the help everyone,

Neil


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