Re: [Histonet] buffer rinses

From:Rene J Buesa

  When I did IHC manually I used to rinse in the following way:
  In a glass container with FRESH (unused) PBS buffer, there was a slide holder; I removed the slides from the wet chamber after the given incubation period was completed and transferred the slides (one at a time) to the slides holders. Once all were there, I just moved the holder with the slides several times (5-7) and that was it.
  Took the slides out (one at a time), wipe them back the slides and around the tissue and added the following reagent.
  While incubating I discarded the used PBS and added fresh one for the next washing. Washing was OK without background and strong signal. If you want you could have 2 containers with PBS and after washing in the first could go to the second. Washing never took more than 1 minutes if 2 washing containers were used.
  Hope this will help you.
  René J.

louise renton  wrote:
  Hi all,

How long and/or vigorous should buffer rinses between immunohisto
steps be(ffpe 4um sections)? I am rinsing 5mins with agitation in PBS
+0.05% Tween. Is this too long? I am concerned that I am washing away
signal yet if I use PBS alone there is overwhelming background and
general muckiness

Louise Renton
Bone Research Unit
University of the Witwatersrand
South Africa
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