MONOCLONAL-ANTIBODIES THAT RECOGNIZE ANTIGENS IN HUMAN OSTEOSARCOMA CELLS AND NORMAL FETAL OSTEOBLASTS

Four monoclonal antibodies (HOB-1-4) that react against human osteosar coma cells (Saos-2) and human fetal osteoblasts in situ were developed by immunizing mice with Saos-2 cells. HOB-1 (IgG1, k) and HOB-2 (IgM, k) stain cytoplasmic antigens in Saos-2 cells, human authentic osteob lasts and occasional cells in liver, spleen and, in the instance of HO B-1, kidney and adrenal gland. On Western blotting of Saos-2 cell lysa tes, HOB-1 recognizes a single protein species of M(r) 59 000, while H OB-2 reacts with a different species of M(r) 57 000. HOB-3 (IgG1, k) s tains a cell membrane associated antigen in Saos-2 cells and osteoblas ts in situ. The reaction pattern of this antibody is virtually identic al to that seen in alkaline phosphatase (ALP)-positive cells in all or gans examined immunohistochemically, except for intestinal epithelium. Both immunoblotting and immunoprecipitation analyses confirm that the antigen detected by HOB-3 is ALP. HOB-4 (IgG1, k) reacts weakly again st Saos-2 cells cultured under standard conditions, but strongly follo wing the exposure of the cells to the secretory inhibitor monensin. On frozen section screening, this antibody reacts preferentially with th e collagenous extracellular matrix not only of the periosteum but of o ther tissues and organs as well. The precise identity of the HOB-4 ant igen remains to be established. We suggest that these four monoclonal reagents will be useful adjuncts in characterizing the osteoblastic ph enotype.