Gayle, I would be interested in the articles as well. I had a similar experience: nerve samples from rats were being shipped to me in K2 (which I had previously sent to them). I specifically said not to freeze the samples; they shipped them on dry ice! I was just doing one micron sectioning, but they were useless.
Thanks! Peggy -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of gayle callis Sent: Friday, May 20, 2011 12:27 PM To: 'Histonet' Subject: [Histonet] Re: Accidental freezing of fish samples You wrote: Probably the bone staining will be OK - how about sending the student to the arctic circle for a few days in winter? - that may be a good lesson about not freezing tissues. Just kidding :-) On Fri, May 20, 2011 at 9:24 AM, Tora Bardal <tora.bardal <http://lists.utsouthwestern.edu/mailman/listinfo/histonet> <@t> bio.ntnu.no>wrote: > One of our students accidentally put his samples in the freezer after formaldehyde/glutaraldehyde/PBS fixation. The samples (fish very early stage, of course rare species) were meant for LM/EM morphology studies. Good morphology is out, but is there a chance > he can do cartilage/bone staining? Or do we need to send him abroad for new sampling at the next spawning time? > Tora Bardal > Senior Engineer > NTNU Center of Fisheries and Aquaculture I was amused by Louise's reply. Your comment "good morphology is out" is correct, especially for EM where bone and cartilage may NOT be optimal after a freeze/ thaw. EM requires optimal fixation and handling to have the best results. There will probably be too much freezing artifact damage (large water ice crystal formation) in the EM samples. However, if doing light microscopy and not worried about soft tissues and with paraffin processing , the bone and cartilage may be ok, but not entirely be ideal ......individual bone and cartilage cells, along with soft tissues, may still show the effects of freezing artifact. These effects may be seen more in cartilage with its higher water content, but give the LM samples a try. You may salvage part of the study. Then send the student out for samples and insist he/she reads up on and understand the damage freezing/thawing can have on fixed samples. I suggest, in lieu of sending your student to the Arctic, that he/she reads the excellent discussion of freezing artifact by Charles Scouten (Myneurolab website) , titled Tips and Techniques: Freezing Biological Samples to prevent this from happening again. A supporting publication, cited by Scouten, is Jongebloed, W.L., Stokroos, D., Kalicharan, D., and Van der Want, J.J.L. Is Cryopreservation Superior Over Tannic Acid/Arginine/Osmium Tetroxide non-Coating Preparation in Field Emission Scanning Electron Microscopy. Scanning Microscopy 13: 93-109, 1999. This paper might help clarify what damage occurs for SEM even though you might be doing TEM (?). I will be happy to send the article in a separate email if you want it. It would be interesting to have follow up on your results, so keep us posted. Good luck Gayle Callis HTL/HT/MT(ASCP) _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet The information in this e-mail is intended only for the person to whom it is addressed. If you believe this e-mail was sent to you in error and the e-mail contains patient information, please contact the Partners Compliance HelpLine at http://www.partners.org/complianceline . If the e-mail was sent to you in error but does not contain patient information, please contact the sender and properly dispose of the e-mail. _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet