The nuclear receptor Rev-erb beta is a potent transcriptional factor whose functional study has been limited by the lack of suitable antibodies against it. To better understand Rev-erb beta' s biological roles, we generated five hybridoma cell lines secreting antibodies against human Rev-erb beta in mice immunized with the purified, prokaryotically expressed recombinant Rev-erb beta-6His fusion protein. Using Western blotting and immunofluorescence analyses, all the five monoclonal antibodies (MAbs) showed strong immunoreactivity to both prokaryotically and eukaryotically expressed recombinant Rev-erb beta. An immunoprecipitation study showed that all five monoclonal antibodies against Rev-erb beta were able to pull down the recombinant Rev-erb beta-Flag protein, but only one of the MAbs against Rev-erb beta, 37H8, could pull down the endogenous Rev-erb beta protein. Furthermore, domain specificity of these MAbs was characterized. Due to the high similarities between Rev-erb alpha and Rev-erb beta in the C and E domains, those C and E domain-specific anti-Rev-erb beta antibodies can react with human Rev-erb alpha as well. The MAbs produced in the study will provide a valuable tool for investigating the function of Rev-erb beta.