RE: [Histonet] tricks about antibody generated in human on human tissue for IHC
Hi Carole, While I have no direct experience with human on human IHC, my first thought would be to biotinylate your primary antibody and then you would omit the anti-human secondary which is the most likely source of your background staining. If you are using fluorescence you could also conjugate your primary with a fluorophore (http://www.invitrogen.com/site/us/en/home/brands/Molecular-Probes/Key-Molecular-Probes-Products/Zenon-Labeling-Technology.html) Brett M. Connolly, Ph.D. Principal Scientist, Imaging Dept. Merck Co., Inc. PO Box 4, WP-44K West Point, PA 19486 brett_conno...@merck.com T- 215-652-2501 F- 215-993-6803 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of carole.chot...@fr.netgrs.com Sent: Friday, January 11, 2013 5:32 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] tricks about antibody generated in human on human tissue for IHC Dear all, I work in an histopathology department and we intend to do immunohistochemistry with an antibody generated in human on human tissues. So far, we don't have any experience with that. So I was wondering if somebody would have some feedback to prevent strong background generated by this human antibody on human tissue. We are planning in a first place to decrease the concentration of the secondary antibody Donkey@human which is recommended at 1/200 by Jackson, using it at 1/500 for example. But if anybody would have some experience with this kind of experiments, their tricks would be more than welcome. Best Regards, Carole Chotard Carole Chotard, Ph.D. Pharmacology Research Scientist Molecular Pharmacology and Pathophysiology Department IDRS 125 Chemin de Ronde 78290 Croissy-sur-Seine France Phone : +33 1 55 72 29 48 Fax : +33 1 55 72 25 96 carole.chot...@fr.netgrs.com ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet Notice: This e-mail message, together with any attachments, contains information of Merck Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates Direct contact information for affiliates is available at http://www.merck.com/contact/contacts.html) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] tricks about antibody generated in human on human tissue for IHC
About 20 years ago in my lab they were developing an anti-human melanoma antibody that we used to test on human melanoma (+) cases and we approached more or less in the way you suggest and were able to eliminate background. You just have to start testing, keep good records of what you do and your results so you can twink your procedure. René J. From: carole.chot...@fr.netgrs.com carole.chot...@fr.netgrs.com To: histonet@lists.utsouthwestern.edu Sent: Friday, January 11, 2013 5:31 AM Subject: [Histonet] tricks about antibody generated in human on human tissue for IHC Dear all, I work in an histopathology department and we intend to do immunohistochemistry with an antibody generated in human on human tissues. So far, we don't have any experience with that. So I was wondering if somebody would have some feedback to prevent strong background generated by this human antibody on human tissue. We are planning in a first place to decrease the concentration of the secondary antibody Donkey@human which is recommended at 1/200 by Jackson, using it at 1/500 for example. But if anybody would have some experience with this kind of experiments, their tricks would be more than welcome. Best Regards, Carole Chotard Carole Chotard, Ph.D. Pharmacology Research Scientist Molecular Pharmacology and Pathophysiology Department IDRS 125 Chemin de Ronde 78290 Croissy-sur-Seine France Phone : +33 1 55 72 29 48 Fax : +33 1 55 72 25 96 carole.chot...@fr.netgrs.com ___ 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
RE: [Histonet] tricks about antibody generated in human on human tissue for IHC
Carole, Look at Golden Bridge International Labs. They have a human-on-human detection system. http://gbi-inc.com/ I've worked with them before and they have excellent products. They also supply some products to other vendors that are rebadged. Tim Morken Supervisor, Electron Microscopy/Neuromuscular Special Studies Department of Pathology UC San Francisco Medical Center -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of carole.chot...@fr.netgrs.com Sent: Friday, January 11, 2013 2:32 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] tricks about antibody generated in human on human tissue for IHC Dear all, I work in an histopathology department and we intend to do immunohistochemistry with an antibody generated in human on human tissues. So far, we don't have any experience with that. So I was wondering if somebody would have some feedback to prevent strong background generated by this human antibody on human tissue. We are planning in a first place to decrease the concentration of the secondary antibody Donkey@human which is recommended at 1/200 by Jackson, using it at 1/500 for example. But if anybody would have some experience with this kind of experiments, their tricks would be more than welcome. Best Regards, Carole Chotard Carole Chotard, Ph.D. Pharmacology Research Scientist Molecular Pharmacology and Pathophysiology Department IDRS 125 Chemin de Ronde 78290 Croissy-sur-Seine France Phone : +33 1 55 72 29 48 Fax : +33 1 55 72 25 96 carole.chot...@fr.netgrs.com ___ 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
