Hi Nicole, I do LFB with IHC on FFPE human brain but there are a few differences in our protocols. I have stained LFB with Neurofilament and with S-100 but never with Ki-67. I also use DAB as my chromagen and differentiate the LFB with dilute Lithium Carbonate. I'm not familiar with the VIP chromagen you mention, so I'm just guessing here (Perhaps somebody on histonet can fill in that blank for us). If VIP chromagen is insoluble in alcohol/xylene, you will likely be fine. I do reduce my staining time in LFB a little so that it doesn't overpower the IHC. You may want to try an LBF protocol with lithium carbonate. It won't hurt IHC at all. One other hint: LFB before IHC doesn't work for me. When I perform antigen retrieval, all of my LFB is lost. If you haven't tried it yet with your current reagents, that would be my first step. If you KNOW that VIP will be removed, I'd try to "borrow" some DAB from a neighbor.
Good luck and don't hesitate to shoot me an email if you have any additional questions. Jennifer L. Hofecker HT(ASCP) Vanderbilt University Medical Center Division of Neuropathology Nashville, TN ph 615.343.0083 fax 615.343.7089 -----Original Message----- From: Patten, Nicole (NIH/NIAAA) [F] [mailto:patte...@mail.nih.gov] Sent: Wednesday, August 26, 2009 10:03 AM To: 'histonet@lists.utsouthwestern.edu' Subject: [Histonet] Luxol Fast Blue with IHC Hello- Has anyone tried to counterstain an IHC stain with Luxol Fast Blue? I am worried that the LFB destain will remove some of the IHC. I use the VIP solution from Vector as the chromogen for IHC instead of DAB and I destain Luxol Fast Blue with Hydroquinone/Sodium Sulfite (in addition to the ethanol steps). I'm staining for Ki-67 and I'm doing this on FFPE human brain sections. I haven't found anything in the Histonet archives on this topic. Any advice would be greatly appreciated. Thanks in advance! -Nicole, Post-bacccalaureate Fellow, NIH _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet