I have tried a blocking solution of 2% BSA and 0.2% triton-X for an hour. I have also tried increasing the BSA to 5% and doubled my blocking incubation time. I am also doing a 30 minute block in 0.1M glycine to account for aldehyde groups in the OCT. My primary antibody is SV-2 and I have tried both Alexafluor 488 and 350. I think it may be an autofluorescence issue but I'm not entirely sure. My other thought is maybe over-permeablization? The protocol calls for 7min in ice cold methanol.