The rise of antimicrobial resistance as a serious global health problem has generated the need for developing nonantibiotic-based therapeutic approaches such as antivirulence strategies, novel vaccines, and host-directed therapies and their widespread use in clinical practice. There is a critical need to develop alternative therapeutic strategies which can disarm bacteria and induce novel clearance mechanisms mediated by the host immune system. In Gram-negative bacteria Pseudomonas aeruginosa, quorum sensing coordinates bacterial behavior as a population and controls the expression of virulence factors associated with pathogenesis. Therefore, blocking bacterial communication may be an attractive strategy for infection control, and our laboratory has developed monoclonal antibodies (mAbs) that can specifically recognize homoserine lactones (HSLs) which control the LuxI/LuxR based QS system in P. aeruginosa. Using sheep immunization and recombinant antibody technology, a panel of sheep-mouse chimeric mAbs were generated which recognized HSL compounds with high sensitivity (nanomolar range) and cross-reactivity. Their therapeutic efficacy was demonstrated in a mouse model of infection, where groups treated with HSL mAbs were protected from lethal Pseudomonas lung infection, prolonging their survival compared to control groups. In addition, the high sensitivity of HSL mAbs makes them ideal for developing immunoassay-based diagnostic test systems for detecting the presence of specific markers of infection (HSLs) in bodily fluids.