Distribution of 65Zn labeled alpha-fetoprotein during proliferation of the BW7756 murine hepatoma

The radiolabeling of alpha-fetoprotein (AFP) with 65Zn for the determination of its biodistribution was studied in mice bearing the BW7756 murine hepatoma as compared to that found with normal mice. AFP is an oncofetal protein of about 70,000 daltons associated with pregnancy and certain cancers (e.g., hepatoma). The AFP was purified from mouse amniotic fluid (MAF) using polyacrylamide gel electrophoresis (PAGE) and higher performance liquid chromatography (HPLC). The biological activity of AFP was maintained through the separation procedures and the purity was determined using double immunodiffusion (DID), immunoelectrophoresis (IEP) and sodium dodecyl sulfate electrophoresis (SDS). The labeling procedures included removal of intrinsic metal with EDTA, incubation with radiotracer (65Zn) and buffer, followed by removal of unbound 65Zn using gel filtration chromatography. The results correlated well with Zn fluctuations recorded by other techniques (RIXRF, radiotracer 65Zn). Large amounts of 65Zn-AFP were localized in the liver, spleen and tumor with significant elevations above normal in the log growth phase (day 14-18). 65Zn-AFP levels in the skin, pancreas, brain and thyroid decreased as the tumor mass increased. Tumor 65Zn-AFP uptake increased with time but leveled off in the late log phase (day 21) due to tumor necrosis. In light of the results of this investigation, and previous work stating that AFP binds Zn with a higher affinity than does albumin, it is suggestive that the Zn fluctuations observed in the earlier hepatoma studies were due to the in vivo binding of Zn to AFP. These results confirm the thesis that intrinsic labeling (replacement of naturally bound ligands with radioactive analogs) does not alter the biochemical integrity as non-intrinsic labeling (e.g., Iodine) may