Metallo- β-lactamase (MBL) is an enzyme that makes bacteria resistant to a broad range of beta-lactam antibiotics. These include the antibiotics of the Carbapenem family, which are a mainstay for the treatment of antibiotic-resistant bacterial infections. The emergence of MBL in Gram negative bacilli (GNB) is becoming a therapeutic challenge worldwide. Detection of MBL is also a challenge for routine microbiology laboratories, since there are no standardized methods for MBL detection. The aims of this study were to know the prevalence of MBL production in various Gram negative bacilli, to evaluate different phenotypic methods to detect MBL production and to find out antibiotic sensitivity profile of MBL producing gram negative bacilli. (Nirav et al., 2011) A total of 100 clinical isolates of GNB were subjected to antibiotic susceptibility testing. Ertapenem resistant clinical isolates were taken as positive for MBL screening. Three different phenotypic methods were used to confirm the MBL production: Ertapenem (ETP)-EDTA combined disc test, EDTA disc potentiation using Ceftazidime and Modified Hodge Test. Out of 100 clinical isolates of GNB, 48 isolates were resistant to Ertapenem. These 48 isolates were considered screening positive and further tested for MBL production by three different methods. 30 isolates were MBL positive by ETP-EDTA combined disc test and 23 isolates were MBL positive by EDTA disc potentiation using Ceftazidime and 19 isolates were positive by Modified Hodge test. In this study, 58.62% species of GNNF, 51.35% of E.coli, 41.67% of Klebsiella pneumonia and 20% of Proteus mirabilis were MBL positive. The detection of MBL-producing isolates is crucial in GNB isolates and ETP-EDTA combined disc test is the most effective method.