Although I tried to perform western blotting with protein extract from RBCs, I could not detect the interest of protein due to red background (maybe, heme or hemoglobin ) all over the lane. I could remove the background largely with TBST of high NaCl concentration, but I could not observe target protein.

Of course, I performed WB with positive control at the same time, and I confirmed that antibody against target protein worked and TBST of high NaCl concentration washed away the protein.

Considering the condition above, I suppose there is a reason below.

1. It is too many protein to perform WB, due to inaccurate measurement of protein concentration.

That is why I ask the question. Could you give me a hand to answer my question or to tell me substitutes or possible causes?