I would use good charged slides like VWR Superfrost Plus or Fisher + slides and try to cut the sections thinner, sections thicker than 4-6 microns fall off easier. It is also important to airdry slides completely before baking them, we bake at 60dc for an hour to overnight in an oven after airdrying. For ish we do it manually on a hot plate and put a glass coverslip over the section to keep it from drying out during high temp long incubations required for ISH.
Patsy Ruegg, HT(ASCP)QIHC Ruegg IHC Consulting, LLC 40864 Arkansas Ave Bennett, CO 80102 Phone: 303-644-4538 Fax: 720-859-4110 pru...@ihctech.net -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Therese C. Thinnes Sent: Friday, August 10, 2012 12:05 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] sections falling off in day 2 ISH (Megha Kumar) Hi Megha, Not sure why your sections are falling off. When I did ISH I used Colorfrost Plus Slides. I would cut at 4 microns-mouse or human. I would dry them at room temp a few hours followed by 55 C for an hour. I would use the slides the next day or next week, never the same day. On day 2 my SSC was 60 C and I used a stir bar for 2 hours on slow stirring. Rarely did I lose any sections. Terri Thinnes Research Assistant The Scripps Research Institute La Jolla, CA Message: 13 Date: Fri, 10 Aug 2012 14:40:30 +0530 From: Megha Kumar <meg...@g.clemson.edu> Subject: [Histonet] sections falling off in day 2 ISH To: histonet@lists.utsouthwestern.edu Message-ID: <cagnji_vjcmlodt3f1zqdk7b79xn3nzcd7ukdigboanbdybg...@mail.gmail.com> Content-Type: text/plain; charset=ISO-8859-1 Hello everyone! When I subject murine skin and intestine paraffin sections (7-10microns) to ISH protocol, the sections fall off on day 2 when I add the SSC. The sections are taken on ploy lysine coated slides and remain on the slides when I do other protocols such as antibody staining. Can anyone suggest me why this is happening? How to prevent the slides from falling off on day 2? Thank you for all the help! regards Megha * _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet