Production and Characterization of Antibodies Against Human Tyrosinase

Abstract

Proteins mapping at different loci are involved in melanogenesis and share several characteristic structural features (b locus, c locus, and slaty locus products). We describe a method to produce specific antibodies against human tyrosinase, the product of the c locus. Mouse L cells transfected with a human tyrosinase cDNA were used to generate antibodies by immunization of syngeneic C3H mice. These antibodies were able to precipitate the tyrosinase glycoprotein from both melanocytic cells and transfectants expressing tyrosinase. In contrast, transfectants expressing the related but distinct b locus protein (gp75 or TRP-1) did not react with these antibodies. In most cases, tyrosinase enzymatic activity could be precipitated and recovered in immune complexes, but one antibody response blocked tyrosinase activity. Immunostaining with anti-tyrosinase antibodies revealed an intracellular granular pattern in tyrosinase transfectants and melanocytic cells, but not transfectants expressing the b locus protein. This approach provides a general method to produce specific antibodies against tyrosinase, other members of the tyrosinase family of proteins, and potentially any other differentiation antigen.