[Histonet] Re: Wrinkles on Glutaraldehyde Fixed Tissue
rleyva <@t> ucsd.edu
Sat May 10 01:26:50 CDT 2014
Your statement makes sense‹glutaraldehyde is quick to fix but slow to
penetrate, and I realized that when comparing the tissue fixation time
frames I used (24h and 48h or more). The tissue that was fixed for 24h is
very hard to spread in the water bath; on the other hand, that which was
in the fixative 48h or longer spread out nicely. Hence, the 24h tissue may
have gotten underfixed, making it difficult to spread and become
On 5/8/14 6:17 PM, "Tony Reilly" <Tony.Reilly <@t> health.qld.gov.au> wrote:
>The problem with Glutaraldehyde is not its fixation but it's penetration
>rate. It is very slow to penetrate which is why EM tissue specimens must
>be diced to such small pieces. If you want to use Glut you will need to
>cut the tissue very thin and use the extended fixation time.
>Tony Reilly B.App.Sc, M.Sc
>Pathology Queensland PAH
>Health Services Support Agency| Department of Health
>Building 15, Level 1,
>199 Ipswich Road
>WOOLLOONGABBA Queensland 4102
>Ph: 07 3176 2412
>Fax: 07 3176 2930
>Email: tony.reillyi <@t> health.qld.gov.au | www.health.qld.gov.au
>From: histonet-bounces <@t> lists.utsouthwestern.edu
>[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Ricardo
>Sent: Friday, 9 May 2014 4:16 AM
>To: Tony Reilly; Bea DeBrosse-Serra; histonet <@t> lists.utsouthwestern.edu
>Subject: [Histonet] Re: Wrinkles on Glutaraldehyde Fixed Tissue
>Thank you for your response. The reason to 4% Glut use is just a PI
>request (really a matter of preference), and it is only on tissue that
>will not be immunostained. The ocular tissue I work with is rabbit. One
>thing I noticed is that the tissue that is fixed for 24h takes longer to
>become wrinkle-free, whereas the 48h (or more) fixed tissue seems to
>spread out in the waterbath with better consistency.
>I am aware of the use of Davidson's<in fact you are the one who so
>patiently gave me recommendations on it, and for that I am grateful.
>Nonetheless, something I'd encounter with Davidson's each time is that
>tissue would shrink a bit too much after processing (all of my
>reagents<EtOH, Xylenes, and Wax are set for about 1h20min each); I tried
>decreasing the processing time but the issue remains. Perhaps the
>processing time needs to be increased on my end to get better results.
>On 5/7/14 7:13 AM, "Bea DeBrosse-Serra" <BDeBrosse-Serra <@t> isisph.com>
>>Why do you use glutaraldehyde for eyes? Davidson's is a much better
>>fixative for eyes. With Davidson's you have two fast penetrating
>>ingredients and one slow acting fixative. Glut is only a fast fixative.
>>Besides, the best institutions who deal with eyes only use Davidson's.
>>Just my thoughts.
>>Beatrice DeBrosse-Serra HT(ASCP)QIHC
>>Antisense Drug Discovery
>>2855 Gazelle Ct.
>>Carlsbad, CA 92010
>>From: histonet-bounces <@t> lists.utsouthwestern.edu
>>[mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of Ricardo
>>Sent: Tuesday, May 06, 2014 11:38 AM
>>To: histonet <@t> lists.utsouthwestern.edu
>>Subject: [Histonet] Wrinkles on Glutaraldehyde Fixed Tissue
>>I am having an issue getting rid of wrinkles on sections from 4%
>>glutaraldehyde fixed paraffin embedded ocular tissue. The laboratory's
>>tissue of interest is the retina, and I use glut to avoid retinal
>>detachment (this is mainly on tissue that won't be immunostained). I
>>normally section at 5µm, and fix the tissue 1 or 2 days. I set the
>>water bath at a temperature that is 5-10 C below the wax melting point.
>>I have tried adding ethyl alcohol to the water and the wrinkles
>>remain-or takes very long for most of them to be gone. I have tried
>>placing the sections in a 1:15 mix of ethanol and water-and have also
>>tried 30% ethanol in water, both at RT, before placing the sections on
>>the water bath; when I do this, the sections separate abruptly (once in
>>the water bath) from the slide I use for transferring the tissue,
>>causing tissue layer separation.
>>When I work with 10% formalin, the section wrinkles disappear nicely in
>>the water bath that contains water and a bit ethanol. Nonetheless, when
>>it comes to 4% glut it just does not work.
>>Any input in troubleshooting this is greatly appreciated.
>>rleyva <@t> ucsd.edu
>>UCSD Shiley Eye Center
>>Histonet mailing list
>>Histonet <@t> lists.utsouthwestern.edu
>Histonet mailing list
>Histonet <@t> lists.utsouthwestern.edu
>This email, including any attachments sent with it, is confidential and
>for the sole use of the intended recipient(s). This confidentiality is
>not waived or lost, if you receive it and you are not the intended
>recipient(s), or if it is transmitted/received in error.
>Any unauthorised use, alteration, disclosure, distribution or review of
>this email is strictly prohibited. The information contained in this
>email, including any attachment sent with it, may be subject to a
>statutory duty of confidentiality if it relates to health service matters.
>If you are not the intended recipient(s), or if you have received this
>email in error, you are asked to immediately notify the sender by
>telephone collect on Australia +61 1800 198 175 or by return email. You
>should also delete this email, and any copies, from your computer system
>network and destroy any hard copies produced.
>If not an intended recipient of this email, you must not copy, distribute
>or take any action(s) that relies on it; any form of disclosure,
>modification, distribution and/or publication of this email is also
>Although Queensland Health takes all reasonable steps to ensure this
>email does not contain malicious software, Queensland Health does not
>accept responsibility for the consequences if any person's computer
>inadvertently suffers any disruption to services, loss of information,
>harm or is infected with a virus, other malicious computer programme or
>code that may occur as a consequence of receiving this email.
>Unless stated otherwise, this email represents only the views of the
>sender and not the views of the Queensland Government.
More information about the Histonet