Re: [Histonet] Incubation of Ab more then 1 hour in autostainer
We used to do overnight in our DAKO Autostainer many years ago, and we kept many containers of dH20 in the incubator to make a humid chamber. Seemed to work well - although we realized overnight was overkill and 1-2 h was sufficient. In fact, given this we started to always put humid chambers inside for all incubations, short or long, as even after 1h we noticed some edge drying on the sections. It improved the quality and reliability of the sections. Andrea --- On Thu, 3/18/10, Margaryan, Naira wrote: From: Margaryan, Naira Subject: [Histonet] Incubation of Ab more then 1 hour in autostainer To: "histonet@lists.utsouthwestern.edu" Date: Thursday, March 18, 2010, 7:38 PM Hi Histonetters! I have a stupid question but I Have to ask. Does anyone perform an Incubation of Ab that required more then 1 hour (2-3 hours) in autostainer? Does autostainer keep slides wet or slides sometimes are getting dry? I know that slides should not be dry in any step of IHC. Thank you very much! Naira ___ 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] Incubation of Ab more then 1 hour in autostainer
Dear Histonetters, Thanks everyone for several good advices. I'll try all suggestions: increase volume from 600ul to 800ul, add more primary antibody step and put hot water dishes inside of machine to keep humidity as high as possible. Hopefully it will work... Many thanks, Naira -Original Message- From: Liz Chlipala [mailto:l...@premierlab.com] Sent: Thursday, March 18, 2010 3:05 PM To: Margaryan, Naira; histonet@lists.utsouthwestern.edu Subject: RE: [Histonet] Incubation of Ab more then 1 hour in autostainer We have done up to an hour but not longer, if I were you I would have two antibody steps and just to add some more reagent after an hour or 1.5 hours Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC PO Box 18592 Boulder, Colorado 80308 office (303) 682-3949 fax (303) 682-9060 www.premierlab.com Ship to Address: 1567 Skyway Drive, Unit E Longmont, Colorado 80504 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Margaryan, Naira Sent: Thursday, March 18, 2010 1:39 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Incubation of Ab more then 1 hour in autostainer Importance: High Hi Histonetters! I have a stupid question but I Have to ask. Does anyone perform an Incubation of Ab that required more then 1 hour (2-3 hours) in autostainer? Does autostainer keep slides wet or slides sometimes are getting dry? I know that slides should not be dry in any step of IHC. Thank you very much! Naira ___ 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: SPAM-LOW: RE: [Histonet] Incubation of Ab more then 1 hour in autostainer
Yea I don't do incubations for longer than one hour on the autostainer and I do put thin trays of water under the slides and on the sides of the machine (it is very dry in Colorado), but I can think of a way you could do longer incubations if you were around, put a coverslip or parafilm over the section after the ab has been applied, you would just have to be there to take it off before the machine goes on to the next steps. This is one area the Leica Bond with coverplates and the Ventana with liq coverslip has over the Dako. I do 2 hour hybridization at 37dc for ish on the bond with the coverplates and the sections do not dry out. Regards, Patsy Patsy Ruegg, HT(ASCP)QIHC IHCtech 12635 Montview Blvd. Ste.215 Aurora, CO 80045 720-859-4060 fax 720-859-4110 www.ihctech.net www.ihcrg.org -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Liz Chlipala Sent: Thursday, March 18, 2010 2:05 PM To: Margaryan, Naira; histonet@lists.utsouthwestern.edu Subject: SPAM-LOW: RE: [Histonet] Incubation of Ab more then 1 hour in autostainer We have done up to an hour but not longer, if I were you I would have two antibody steps and just to add some more reagent after an hour or 1.5 hours Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC PO Box 18592 Boulder, Colorado 80308 office (303) 682-3949 fax (303) 682-9060 www.premierlab.com Ship to Address: 1567 Skyway Drive, Unit E Longmont, Colorado 80504 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Margaryan, Naira Sent: Thursday, March 18, 2010 1:39 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Incubation of Ab more then 1 hour in autostainer Importance: High Hi Histonetters! I have a stupid question but I Have to ask. Does anyone perform an Incubation of Ab that required more then 1 hour (2-3 hours) in autostainer? Does autostainer keep slides wet or slides sometimes are getting dry? I know that slides should not be dry in any step of IHC. Thank you very much! Naira ___ 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 ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
Re: [Histonet] Incubation of Ab more then 1 hour in autostainer
Hi Naira, We routinely do a 2 hour incubation in primary, but we make sure that more antibody is on the slide than would be necessary for most other reagents (500-600 ul). We have not had any issues of drying. If you are concerned about it, just make it 2 antibody steps instead of 1 on the machine. Kim Kim Merriam, MA, HT(ASCP)QIHC Cambridge, MA From: "Margaryan, Naira" To: "histonet@lists.utsouthwestern.edu" Sent: Thu, March 18, 2010 3:38:33 PM Subject: [Histonet] Incubation of Ab more then 1 hour in autostainer Hi Histonetters! I have a stupid question but I Have to ask. Does anyone perform an Incubation of Ab that required more then 1 hour (2-3 hours) in autostainer? Does autostainer keep slides wet or slides sometimes are getting dry? I know that slides should not be dry in any step of IHC. Thank you very much! Naira ___ 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] Incubation of Ab more then 1 hour in autostainer
We have done up to an hour but not longer, if I were you I would have two antibody steps and just to add some more reagent after an hour or 1.5 hours Liz Elizabeth A. Chlipala, BS, HTL(ASCP)QIHC Manager Premier Laboratory, LLC PO Box 18592 Boulder, Colorado 80308 office (303) 682-3949 fax (303) 682-9060 www.premierlab.com Ship to Address: 1567 Skyway Drive, Unit E Longmont, Colorado 80504 -Original Message- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Margaryan, Naira Sent: Thursday, March 18, 2010 1:39 PM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Incubation of Ab more then 1 hour in autostainer Importance: High Hi Histonetters! I have a stupid question but I Have to ask. Does anyone perform an Incubation of Ab that required more then 1 hour (2-3 hours) in autostainer? Does autostainer keep slides wet or slides sometimes are getting dry? I know that slides should not be dry in any step of IHC. Thank you very much! Naira ___ 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
