New methods are needed to eradicate or prevent Chlamydia trachomatis infections. Blockade of epithelial membrane protein 2 (EMP2) by genetic silencing or neutralizing polyclonal antibody reduced chlamydial infectivity in vitro. This study tests the prediction that recombinant anti-EMP2 diabody could reduce early chlamydial infection of the genital tract in vivo. In a murine infection model, pretreatment with anti-EMP2 diabody, as compared with control diabody, significantly reduced bacterial load, tissue production of inflammatory cytokines, recruitment of polymorphonuclear leukocytes, and local tissue inflammation. These findings support EMP2 as a potential preventative and therapeutic target for genital chlamydial infection.