Re: [Histonet] Re: Buffer.

From:John Kiernan

   PIPES-buffered  glutaraldehyde  has  been used for subsequent electron
   micr=  oscopy  since  the  early  1970s but it has been shown to cause
   artifacts    -   =   intracellular   vesicles   and   myelin   figures
   - when  used  for  perfu=  sion  fixation of the brain. See
   Schultz  RL=  &  Wagner DO (1986)
   Membrane  alterations  in  cerebral cortex when= using PIPES buffer.
   J.  Neurocytol.  1= 5:
   461-469.  The  authors  attributed  these  art= ifacts,
   which  potentially  could  be mistaken for pathological changes,= to
   the  low toxicity of PIPES. Apparently cacodylate or phosphate buff   er  prevented  the  metabolic changes that caused the artifacts during
   the  =  early  phase  of  fixation, but PIPES allowed the changes to
   occur.  <=  BR> 
The reason for using cacodylate buffer rather tha= n phosphate is that it doesn't precipitate with calcium ions. A Ca s= alt in the fixative improves the immobilization of phospholipids in cell membranes.
John Kiernan
Anatomy, UW= O
London, Canada
= = =
----- Original Messa= ge -----
From: Robert Richmond < gt;
Date: Sunday, November 16, 2008 14:13
Subjec= t: [Histonet] Re: Buffer.
To: histonet@lists.utso

> About the potential toxicity of= arsenic-containing cacodylate buffers:
>= A Histonetter who prefers to remain anonymous notes >>The= arsenic
> discussion comes up on the Microscopy Listserve= r every once in while,
> usually because someone is eith= er pregnant & concerned about exposure
> or has sa= fety officers who freak out over the stuff. It was used in
&g= t; the early days because phosphate buffers can interact with some <= BR>> of the
> fixative combinations and leave pre= cipitates. Also, I'd suspect,
> cacodylate is less= prone to beasties growing in stock solutions over
> time.= The "modern" zwitterionic buffers are used a lot these
&g t; days -
> PIPES & that crowd - if you want to = avoid cacodylate &
> phosphate.<<=
> Those zwitterionic buffers are often referred to as Goo= d's
> buffers or
> Good buffers, since = they were introduced by Norman Good in the 1960's.
> S= ee
> I don't know why they aren't commo= nly used as buffers for
> histologic fixatives.
= >
> Bob Richmond
> Samurai Pathologi= st
> Knoxville TN
> ___ _______________________ 5F ____________________
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