The production and characterization of murine monoclonal antibodies to human Gadd45 raised against a recombinant protein.

Abstract

The production of two different murine monoclonal antibodies to human Gadd45, a protein that is induced in response to DNA damage, is reported. Antibodies were generated in a SJL mouse using a recombinant form of the human Gadd45 protein. Monoclonal antibody 4TCYA1, which recognizes the denatured form of human Gadd45 in Western blots, was selected based upon the recognition of Gadd45 induced by functional p53 in the human myeloid leukemia cell line, ML-1. A second monoclonal antibody, designated 30T.14, immunoprecipitates native human Gadd45 in lysates produced from RKO cells, a colorectal carcinoma cell line that expresses relatively high basal levels of Gadd45, as well as from cell lysates made from ML-1 cells after exposure to ionizing irradiation (IR). Since 4TCYA1 fails to immunoprecipitate Gadd45, and 30T.14 fails to bind to IR-induced Gadd45 in immunoblotting, these two monoclonal antibodies probably recognize different epitopes.