From owner-drosophila@net.bio.net Sun Sep 10 23:00:00 1995 Path: biosci!bcm.tmc.edu!news.msfc.nasa.gov!newsfeed.internetmci.com!howland.reston.ans.net!tank.news.pipex.net!pipex!news.sprintlink.net!in2.uu.net!sparky!nlcnews.nlc.state.ne.us!crcnews.unl.edu!manager From: "Alan C. Christensen, Ph.D." Newsgroups: bionet.drosophila Subject: (no subject) Date: 11 Sep 1995 17:12:25 GMT Organization: University of Nebraska Lines: 22 Message-ID: <431qlp$d9b@crcnis3.unl.edu> NNTP-Posting-Host: bs-achristensen1.unl.edu Mime-Version: 1.0 Content-Type: text/plain; charset=us-ascii Content-Transfer-Encoding: 7bit X-Mailer: Mozilla 1.1N (Macintosh; I; PPC) X-URL: news://crcnews.unl.edu/bionet.drosophila Here's a simple question, which I hope someone has a simple answer to. We are staining whole-mount embryos with an antibody, and then with an HRP-conjugated second antibody. We are getting signal, but there is some background, presumably from endogenous peroxidases. The standard protocol to inactivate peroxidases is to incubate with 3% H2O2 in methanol, just after devitellinizing. However, the antigen is methanol sensitive, so we are hand-peeling the embryos. I've been told that treatment of specimens with 3% H2O2 in aqueous solutions results in violent bubbling and destruction of the specimen. We will be trying a few things, but if anyone has a successful method for inactivating endogenous peroxidase without methanol, please let me know. ***************************************************************** * Alan C. Christensen, Ph.D. "...there are those rare * * School of Biological Sciences characters who study the * * University of Nebraska unknown product of an * * Lincoln, NE 68588-0118 unknown gene. These we * * Phone 402-472-0681 call lunatics." * * FAX 402-472-2083 -Sidney Brenner * ***************************************************************** .