We found that some plasma proteins such as haptoglobin (Mr 200,000) and alpha 1-acid glycoprotein (Mr 40,000) do not bind at all to nitrocellulose membranes in classical transfer conditions. In contrast, these proteins can be clearly identified when a charged nylon membrane is disposed on the anodal side of the nitrocellulose membrane. Several factors which are important in modifying the adherence between proteins and the blotting membrane have been studied. They are not directly related to the molecular weight of the protein considered but also include the presence of organic solvents in the buffer systems, the pore size of the membrane and the degree of reduction and/or glycosylation of the protein. The eventuality of decreased or absent identification of some antigens on nitrocellulose membranes stresses the importance of controls using nylon membranes.