I am honored to have became member of such Community!
I am asking you help because I got a frustrating problem with WB.
I am searching an aromatasis enzyme (p450c17, 55 kDA) on rat brain homogenates but, at this time, I can't get any results. I am following a protocol validated for many other proteins I worked with and I can exclude any major problem durig the procedure at least until the transfer since the Ponceau staining revealed a good transfer, a good run of protein and the marker is well defined as well. However, the weird thing is that when I perform dot-blot I got a positive reaction. I am performing IF on the same brain region in order to see what will happen there, in the meanwhile as it regard WBi have no any idea or solutions on what to do.
I appreciate any suggestions....