Re: [Histonet] What is the best microtome?

[Dr. Michael Gudo via Histonet]

We have Leica microtomes in out lab and they work very very good. The 
components are compatible over several Generations and they work for at least 
more than 15-20 years!
And the best thing is the Waterbath directly attached to the Blade.

Greetings
Michael 

Von meinem iPhone gesendet

> Am 14.09.2017 um 00:18 schrieb Cooper, Brian via Histonet 
> :
> 
> Microm HM325 gets my vote.
> 
> -Original Message-
> From: Dawn Bugge via Histonet [histonet@lists.utsouthwestern.edu]
> Received: Wednesday, 13 Sep 2017, 3:04PM
> To: histonet@lists.utsouthwestern.edu [histonet@lists.utsouthwestern.edu]
> Subject: [Histonet] What is the best microtome? (EXTERNAL EMAIL)
> 
> Hello everyone,
> 
> What is your favorite microtome?  I am helping a lab with purchases and I
> only have experience with the Finesse ME+ and an old Leica.  Any thoughts
> would be great.
> 
> Thanks.
> 
> --
> Dawn R Bugge
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Re: [Histonet] Paraformaldehyde Fixed Tissue

[Michael Gudo via Histonet]

Dear Sandra,
Dear Ana,

in our lab for tissues for which we have enough time for infiltration we fix 
for at least 48 hours or more in NBF or PFA (both with 4 % reactive 
formaldehyde, which is equivalent to 10% „neutral buffered formaline“) and then 
first wash in tap water for a minimum of 1-2 hours, then to 30%, 50%, 60%, 70%, 
80%, 90% 96% (2 times), 2-Propanol (2-times) and then 2-3 time xylene, before 
embedding in paraffine. The times are just a few hours per step or shorter, and 
we have quite good results especially for brain and spinal chord.

Best regards
Michael



> Am 29.08.2017 um 02:06 schrieb Ana Maluenda via Histonet 
> :
> 
> Hi Sandra,
> 
> I haven't myself particularly worked with brain and spinal cord, but majority 
> of my protocols in my old job used fixation in 4% PFA (24 hours at 4-8oC) and 
> routinely process (or transfer to graded EtOH, if not processing immediately).
> However, our routine process didn't include a first step in 10% NBF, since 
> PFA plays the role of fixation. Therefore, after PFA, we would have 70% EtOH, 
> 80% EtOH, 95% ETOH, 2x EtOH the Xylenes and wax [assuming you are referring 
> to FFPE?].
> Also mouse tissue can be small and delicate, so I remember running liver, 
> spleen, kidney and thymus (soft tissues I worked with) in a short cycle 
> (similar to what I would do for biopsies).
> 
> Hope this helps!
> 
> Kind regards,
> 
> Ana
> 
> 
> Ana Maluenda
> Research Assistant
> Atherothrombosis and Vascular Biology Laboratory
> 
> Baker Heart and Diabetes Institute
> 75 Commercial Road, Melbourne VIC 3004
> P (03) 8532 1359 E ana.malue...@baker.edu.au W www.baker.edu.au
> 
> -Original Message-
> From: Sandra Cheasty [mailto:sandra.chea...@wisc.edu]
> Sent: Tuesday, 29 August 2017 1:30 AM
> To: Histonet (histonet@lists.utsouthwestern.edu) 
> 
> Subject: [Histonet] Paraformaldehyde Fixed Tissue
> 
> Hi all,
>   We are having difficulty sectioning mouse tissue, (brain, 
> spinal cord, liver, and spleen), on paraformaldehyde fixed tissue. Has anyone 
> had issues with paraformaldehyde fixed tissue? They were processed routinely, 
> starting in 10% NBF, with other tissues, and we are cutting them at 3u.
> Thank you!
> Sandy
> 
> Sandra J. Cheasty, HT (ASCP)
> Histology & Necropsy Supervisor
> UW-Madison, School of Veterinary Medicine
> 
> 
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Re: [Histonet] Cut FFPE Slides for ISH

[Michael Gudo via Histonet]

Hello Jenniffer,

we store the slides with section at 4-8 °C for many month or even longer and we 
do not have any problems with ISH.

Best regards
Michael



> Am 28.08.2017 um 17:30 schrieb Jennifer Phinney via Histonet 
> :
> 
> Hello Histonet,
> I am hoping someone can point me in the right direction to find some 
> references for how long FFPE slides intended for ISH can be kept before 
> staining.  I have a researcher who swears their slides have to be stained 
> within 24 hours of being cut, which does not sound correct to me.  I've used 
> ACD's RNAscope before and their protocol lists the timeframe as 3 months (so 
> this is a bit of a difference).
> 
> There's a lot of information about fixation times, but I've come up empty 
> handed for cut slides.
> 
> Thanks everyone!
> 
> Jennifer Phinney QIHCCM
> Project Coordinator
> Kansas State University
> Veterinary Diagnostic Lab
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