To identify vitamin B12-binding proteins in egg white, an affinity chromatographic isolation procedure was applied. A fraction tightly adsorbed on vitamin B12 immobilized in agarose was eluted with 1 mM cyanocobalamin, and then separated by isoelectric focusing in sucrose density gradient. Two isolated proteins (or perhaps two forms of the same protein) of isoelectric points 6.2 and 7.2 and the same Mr 90,000 were found capable of detectable vitamin B12 binding. They are probably glycoproteins, each composed of a single polypeptide chain.