I agree most of your times are too long, but you can still get sections if you put water on your ice block and allow the tissue to rehydrate a bit or put a little ammonium hydroxide and water on your ice and sit your blocks to be section in this solution.
-----Original Message----- From: [email protected] [mailto:[email protected]] On Behalf Of Merced M Leiker Sent: Wednesday, November 10, 2010 9:17 AM To: Michael Mashore; [email protected] Subject: Re: [Histonet] Paraffin Tissue Crumbles Having some experience processing and embedding rodent tissues myself (by hand), I would say that you are over-dehydrating the tissues. Try cutting back the alcohol incubation times to 30 min or even 10 min each. Regards, Merced --On Tuesday, November 09, 2010 2:56 PM -0800 Michael Mashore <[email protected]> wrote: > Hello Histonet Users, > > > > I have just started using paraffin and am having many difficulties. > Most of the time my tissue crumbles when sectioning. I have no real > experience in paraffin histology and have been given the task of > becoming proficient by myself, so I am hoping for feedback as to why > my tissue keeps crumbling. The tissue in question has been: skeletal > muscle, cardiac muscle, liver, and brain (all from rat). > > > > The tissue was fixed in 10% neutral buffered formalin for 7 days at > 4°C and then transferred to an automated tissue processor, with the > following > schedule: > > > > 2 hours 70% dehydration alcohol > > 2 hours 80% dehydration alcohol > > 2 hours 95% dehydration alcohol > > 2.5 hours 95% dehydration alcohol > > 2 hours 100% dehydration alcohol > > 2 hours 100% dehydration alcohol > > 2 hours 100% dehydration alcohol > > .5 hour Hemo-De > > .5 hour Hemo-De > > .5 hour Hemo-De > > 1 hour paraffin > > 4 hours paraffin > > > > They were infiltrated for 1 hour without vacuum then embedded. > > The blocks were stored in the freezer before cutting. > > The knife angle was 5°. > > Sections were 5µm thick. > > > > I would appreciate any feedback whatsoever. > > > > Thank you very much. > > > > Michael > > _______________________________________________ > Histonet mailing list > [email protected] > http://lists.utsouthwestern.edu/mailman/listinfo/histonet > Merced M Leiker Research Technician III Cardiovascular Medicine 348 Biomedical Research Building State University of New York at Buffalo 3435 Main St, Buffalo, NY 14214 USA [email protected] 716-829-6118 (Ph) 716-829-2665 (Fx) No trees were harmed in the sending of this email. However, many electrons were severely inconvenienced. _______________________________________________ Histonet mailing list [email protected] http://lists.utsouthwestern.edu/mailman/listinfo/histonet ------------------------------------------------------------------------------ CONFIDENTIALITY NOTICE: The information in this e-mail may be confidential and/or privileged. If you are not the intended recipient or an authorized representative of the intended recipient, you are hereby notified that any review, dissemination, or copying of this e-mail and its attachments, if any, or the information contained herein is prohibited. If you have received this e-mail in error, please immediately notify the sender by return e-mail and delete this e-mail from your computer system. Thank you. ============================================================================== _______________________________________________ Histonet mailing list [email protected] http://lists.utsouthwestern.edu/mailman/listinfo/histonet
