[Histonet] Re:Endogenous biotin blocking
Hello, From: Hobbs, Carl carl.ho...@kcl.ac.uk Subject: [Histonet] Re:Endogenous biotin blocking If you are in a research lab and find that you will need to apply biotin blocking regularly, you can make up your own avidin/biotin solutions if costs are a major factor. To overcome the problem of endogenous biotin I incubated the sections in an avidin solution. The avidin was the avidin-DH form that comes with the Vector Stain ABC kits. I added it to my blocking solution as per the manufactures instructions and incubated it on the sections for 1 hr at room temperature. There after the sections were washed 3x in buffer and then I added biotin to a final concentration of 1 mg/mL to the blocking solution and incubated for 15 minutes at room temperature. After the biotin step the biotinylated probe in blocking solution was added and incubated over night at 4 deg C. This worked very well! Almost all back ground due to the endogenous biotin was quenched. Regards -- Tyrone Genade http://tgenade.freeshell.org email: tgen...@gmail.com tel: +27-84-632-1925 (c) Romans 6:23: The gift of God is eternal life through Christ Jesus our Lord. To find out how to receive this FREE gift visit http://www.alpha.org. ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
[Histonet] Re:Endogenous biotin blocking
Your understanding is correct, imho. I add a no primary control on my stABCpx-DAB run whenever I am testing new tissues and assess any positivity that could be due to endogenous biotin. You will quickly build up a knowledge of those tissues that automatically require biotin - blocking. ( eg liver, kidney) If you are a clinical/registered Lab, you wil of course be required to buy a commercial kit. If you are in a research lab and find that you will need to apply biotin blocking regularly, you can make up your own avidin/biotin solutions if costs are a major factor. I will be interested in other suggestions/approaches. carl ___ Histonet mailing list Histonet@lists.utsouthwestern.edu http://lists.utsouthwestern.edu/mailman/listinfo/histonet
