In Vitro Observations on the Binding of Vitamin B 12 by Serum Proteins and the Effect of Cells on the Process

Summary

In vitro studies of the binding of vitamin B₁₂ in the presence and absence of cells were done. Sera from 3 healthy volunteers were incubated with a trace amount of ⁵⁷CoB¹² and aliquots were incubated with and without autologous peripheral blood cells. At the same time, the sera were incubated in this fashion but without added ⁵⁷CoB₁₂ to determine the effect of the incubation conditions on the UBBC.

The results showed the following: (1) UBBC and its 3 component binding proteins remained constant over 24 hr, indicating that related observations were probably not due to denaturation of proteins. Fall in UBBC after 24 hr was due to decrease, possibly denaturation, of TC II while TC I remained constant; this may reflect the short half-life of TC II and the long half-life of TC I in vivo. (2) In the presence of cells, however, serum UBBC rose with time as a result of increasing TC I and third binder. Both binders rose equally over 24 hr while the cells remained intact, third binder becoming particularly increased thereafter as cell destruction became noticeable. These findings demonstrate that, unlike TC II, TC I and third binder are derived from blood cells. The differing patterns of their increase suggest that the latter 2 proteins are not identical. (3) Trace ⁵⁷CoB₁₂ binds preferentially to TC II and is initially transferred predominantly to cells. Later, transfer occurs to third binder, if present, and to a lesser extent to TC I. It appears, but is not certain, that transfer to TC I and third binder is direct, since it takes place even in the absence of cells. (4) TC II appears to attach to cells whether it is carrying ⁵⁷CoB₁₂ or not.