Cobalamin-binding proteins in human urine: identification and quantitation

The urinary cobalamin (Cbl) binders were examined by gel filtration of concentrated urine on Sephadex G-200, alone and together with immunoglobulins against human Cbl binders. The 11 urine specimens from 10 persons studied had a free Cbl-binding capacity of 0.76 to 30.0 pmol/L (mean 6.6 pmol/L). Gel filtration with buffer preventing association of the Cbl binders gave peaks with elution volumes characteristic of serum transcobalamin and haptocorrin. When concentrated urine specimens were run together with immunoglobulins, the findings confirmed the presence of haptocorrin and transcobalamin and, in addition, of intrinsic factor. The presence of intrinsic factor was confirmed by incubating urine with an ion exchanger gel and by showing that an eluate of the gel reacted with the corresponding immunoglobulin. Part of the urinary intrinsic factor detected by radioimmunoassay was able to bind 57Co-Cbl; the rest appeared to be saturated with endogenous Cbl and possibly partly incapable of binding Cbl. All the free Cbl-binding capacity of urine concentrate could be blocked with adenine cyanocobamide, but only half with cobinamide; cobinamide blocked all the urinary haptocorrin but only part of the transcobalamin and none of the intrinsic factor eluted from the ion exchanger gel.