Incorporation of vitamin B12 into L1210 cells requires the protein binder transcobalamin II (TCII). The process is saturable, follows Michaelis-Menten kinetics (Km = 2.5 X 10(-9) M at 37 degrees C), is both temperature and calcium (K50 - 1 X 10(-6) M) dependent, and is inhibited by apo-TCII, indicating the presence of a TCII specific receptor on the cell membrane. B12 also leaves the cell by a calcium-independent pathway bound to either TCII or to a protein with chromatographic properties similar to those of TCIII. Since intact TCII-B12 can be found in the cytosol and can promote B12 uptake by mitochondria, it is proposed that the B12 released from the cell bound to the TCIII-like protein is derived by mitochondrial processing of incorporated TCII-B12. The slower time course of release of the latter B12 is consistent with this postulate.