Background: This study was undertaken to determine the prevalence of Metallo-β-lactamase (MBL) producing gram negative bacteria in a tertiary care hospital in North India. Metallo-β-lactamases are β-lactamase enzymes with distinct ability to hydrolyze and confer resistance to the carbapenems which are the commonly used drugs for the treatment of infections caused by multidrug resistant gram negative bacteria. Pathogenic bacteria producing Metallo-β-lactamases are responsible for infections in tertiary care hospitals around the world and carbapenem resistance due to these enzymes have been reported over the past few years. Material and Methods: A total of 400 Enterobacteriaceae isolates from various clinical samples were identified and tested for susceptibility to various antibiotics by disc diffusion method. MBL detection was done by combined disc diffusion test and E-test. Result: The isolates included, E.coli 170(42.50%), K. pneumoniae 113(28.25%), Citrobacter spp. 90(22.50%) & Proteus spp. 27(6.25%).Out of the 400 Enterobacteriaceae 77 were carbapenem resistant of these 77 isolates 42 (54.54%) showed an increase in zone size by the combined disc diffusion test and E-test. MBL was detected in 27(30.33%) isolates of Citrobacter spp, 26 (23.0%) K. pneumoniae, 22 (12.94%) E.coli, 2 (7.40%) Proteus spp isolates. Conclusion: The study highlights the problem of MBL mediated resistance, which indicates therapeutic challenge for the clinicians and microbiologists. Simple disc method can be routinely employed to detect these common resistance mechanisms which will reduce the mortality and also spread of such resistant strains. In view of the increasing resistance of gram negative bacilli to carbapenems, rational use of antibiotics needs to be emphasised.