Need some info and insights anyone might have on the biotinylated cytokine kits available for cytokine receptor detection from R&D Systems. We are trying to use their TNF kit to detect TNF-R1 in the mouse (Human TNF signals through TNF-R1 but not TNF-R2 in the mouse). We get a subtle shift, and are looking to modify the protocol to improve performance, but can't get R&D Systems to tell us if their wash buffer has some kind of fixative or special detergent in it (maybe something called BSCO??). Their protocol has no wash step after addition of cytokine and before addition of streptavidin secondary (we already switched from their sAV-FITC to a sAV-APC). We think a wash step here may allow us to increase the amount of biotiylated cytokine we use without increasing our background too much. If anyone can give insights into how you may have optimized these kits, or what might be in the "secret recipe" of R&D's wash buffer (so we can modify), please drop me a line. thanks Keith Bahjat kbahjat@ufl.edu
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