[Histonet] DRGs and Histogel

Esther Peters esther.peters <@t> verizon.net
Mon Feb 22 16:36:39 CST 2010 

   I  have  had  the  exact  same  results, one piece processing well and
   another  in  the  same  cassette  shrinking  and hardening, when using
   1.5-2% low melting point agarose (NuSieve) instead of Histogel.  I was
   thinking  of  changing  to Histogel, but then this thread started last
   summer  and  now I see issues continue.  I would love to know the best
   way to do this also!
   Esther Peters, Ph.D.
   George Mason University
   dusko trajkovic wrote:

I also think that it is strange of the way Histogel processes. I have posted on
 the Histonet previously about this exact problem. I worked with Jennifer Hofec
ker when she was at Vanderbilt U.(sent her my Histogel and she sent me hers) an
d ended up with perfectly processed Histogel blocks at our facility and hers. I
 processed a couple of blocks last week and they were just terrible. No change 
in the processing schedule, or the way Histogel was liquefied (placed in hot wa
ter that was heated in the microwave). Prior to the last two blocks, I must hav
e processed at least a dozen blocks without any problems. There was an incident
 where I placed two histogels in the same cassete. One processed beautifuly and
 the other was all shrunken and dried up.
I do not liquefy the entire tube, rather I scoop out the approximate amount tha
t I need and transfer to another tube to heat up. If there is anyone out there 
in Histoland that has not had any issues with the Histogel, can you please post
 your procedure on liquefying the Histogel, method of cooling/solidifying and p
rocessing schedule? The only thing that I do that is not exact is I do not know
 the temp of my hot water when i place the Histogel to liquefy. I basically hav
e to wait several minutes for the gel to melt and I use it immediately.
Any new information or solution from anyone, would be greatly appreciated.
Thank you
Dusko Trajkovic



________________________________
From: Amy Porter [1]<portera <@t> msu.edu>
To: Andrea Grantham [2]<algranth <@t> email.arizona.edu>; HISTONET [3]<histonet <@t> list
s.utsouthwestern.edu>
Sent: Mon, February 22, 2010 9:01:22 AM
Subject: RE: [Histonet] DRGs

I think it is strange that we are all doing similar techniques and wind up
with different outcomes using the histogel.  I would be curious how many of
us are using the equipment sold with the histogel for warming and cooling
opposed to any of us who don't.  we did not purchase the equipment and I
wonder sometimes if warming the histogel using other means causes some type
of breakdown / and do any of you repeatedly reheat the same tube once it has
been warmed and resolidified??

-----Original Message-----
From: [4]histonet-bounces <@t> lists.utsouthwestern.edu
[[5]mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Andrea
Grantham
Sent: Monday, February 22, 2010 9:41 AM
To: HISTONET
Subject: Re: [Histonet] DRGs
Importance: High

Hi Carol,
I have used histogel for these kinds of samples and also other small,
thin tissues like insect antennae and insect GI tracts and midguts.
Since I get all my projects already fixed in whatever fixative the
investigator chooses, rinsed and placed in 70% ETOH the histogel never
touches formalin. I don't use formalin on my processor but start in
70%. I've never had a problem with the histogel. We just put the
sample in the histogel flat and stand it up (turn 90°) when embedding
in paraffin. I use tissue prep for embedding.
If you don't want to use histogel you could try to put the drg's on GN
Metricel membrane disc filters. We do this with a lot of the samples I
receive, actually I have the investigators or their techs do this. The
tissue sticks to the membrane and orientation is a dream. The membrane
presents no problem when sectioning. You can get it from VWR.

Andi



Andrea Grantham, HT (ASCP)
Senior Research Specialist
University of Arizona
Cell Biology and Anatomy
Histology Service Laboratory
P.O.Box 245044
Tucson, AZ 85724

[6]algranth <@t> email.arizona.edu
Tel: 520.626.4415    Fax: 520.626.2097

"happy slicing and dicing and may all your stains work perfectly" -
Paula Sicurello
P Please consider the environment before printing this email.




On Feb 18, 2010, at 11:23 AM, Barone, Carol wrote:



Histonetter's...we received a "boat-load" of mouse DRGs that had been
prepared in histogel and are cutting...well..not so good.
We normally do DRGs from FS and get beautiful results.

We have used histogel before with other small sample and have never
had
issues...  not sure if it is the Histogel or DRG's (fixed in 10% NBF
and
then transferred to 70% before placed into the histogel).is the
issue..I
seem to remember that histogel requires formalin and wonder if the
transfer to 70% is causing our problem ...but, obviously there is not
much room for error with such tiny- tiny samples and they are already
process and in paraffin?

I am not quite sure how twe can improve the ones that came in
histogel,
and were processed to paraffin a paraffin block....any idea's? any
experience? any anything? Thx- ASAP!

[7]cbarone <@t> nemours.org
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References

   1. mailto:portera <@t> msu.edu
   2. mailto:algranth <@t> email.arizona.edu
   3. mailto:histonet <@t> lists.utsouthwestern.edu
   4. mailto:histonet-bounces <@t> lists.utsouthwestern.edu
   5. mailto:histonet-bounces <@t> lists.utsouthwestern.edu
   6. mailto:algranth <@t> email.arizona.edu
   7. mailto:cbarone <@t> nemours.org
   8. mailto:Histonet <@t> lists.utsouthwestern.edu
   9. http://lists.utsouthwestern.edu/mailman/listinfo/histonet
  10. mailto:Histonet <@t> lists.utsouthwestern.edu
  11. http://lists.utsouthwestern.edu/mailman/listinfo/histonet
  12. mailto:Histonet <@t> lists.utsouthwestern.edu
  13. http://lists.utsouthwestern.edu/mailman/listinfo/histonet
  14. mailto:Histonet <@t> lists.utsouthwestern.edu
  15. http://lists.utsouthwestern.edu/mailman/listinfo/histonet

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