Hi, I am just wondering if anyone has run into this problem before. I am using the licor imager to try to probe for 2 proteins, Girk and D2.

I blocked with the licor blocking buffer 1:1 with PBS. both the primary and the secondary antibody was mixed with this buffer. Primary was incubated overnight at 4 degrees and secondary was 1 h at rt. 

I also made sure to dip the membrane in methanol before, and after the transfer.

Someone mentioned to me that the streaks could be from Ponceau s. I did use ponceau before but not this time, however I used the same antibody mixture again so it could have washed off a little into the antibody mix.  

photo attached