Hi Natalia, Human tissue is naturally autofluorescent. In our lab we do immunos on frozens and quench the AF with chicago blue solution (1g Chicago Sky Blue 6B-Sigma in 99ml MilliQ plus 1ml DMSO) after washing the secondary antibodies with PBS (3x). Just load the tissue with a drop of the CB solution for 15 secs and wash it in PBS again (3x). Mount as usual
D -----Original Message----- From: histonet-boun...@lists.utsouthwestern.edu [mailto:histonet-boun...@lists.utsouthwestern.edu] On Behalf Of Natalia Fernandez Sent: Wednesday, 8 February 2012 2:35 AM To: histonet@lists.utsouthwestern.edu Subject: [Histonet] Immunofluorescence Hi everybody! I have a problem with my experiment. I try to do immunofuorescences in old human brains (parafine sections). First I thought that the too much colocalisation was due to my antibodies (primary or secondary). After several tests (Only 1st antibody, only 2nd one, simple staining, inverse simple staining,..) I saw that the tissue itself shows a lot of fluorescence (without antibodies, and even after a treatment against lipofuscine (solution of Potassium permanganate). So my question is: Do you have the same problem? Do you think that parafine could be the reason of this autofluorescence?! What can I do? Thank you so much for your answers :) Natalia _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet ______________________________________________________________________ This email has been scanned by the Symantec Email Security.cloud service. For more information please visit http://www.symanteccloud.com If you have any question, please contact MCRI IT Helpdesk for further assistance. ______________________________________________________________________ ______________________________________________________________________ This email has been scanned by the Symantec Email Security.cloud service. For more information please visit http://www.symanteccloud.com ______________________________________________________________________ _______________________________________________ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet