EVALUATION OF 5 DIFFERENT METHODS TO PREPARE BACTERIAL EXTRACTS FOR THE IDENTIFICATION OF BETA-LACTAMASES BY ISOLECTRIC FOCUSING

The X-PRESS, osmotic shock, chloroform treatment, lysozyme treatment and ultrasonic disruption methods to release five different plasmid-mediated β-lactamases from Escherichia coli and one chromosomal β-lactamase from Enterobacter cloacae were compared. The main activities of TEM-1, SHV-1, OXA-1, OXA-2, PSE-4 and chromosomal PSE β-lactamases were found at the same isoelectric point irrespective of the method used. However, additional satellite bands were found with TEM-1, OXA-1, OXA-2 and PSE-4 β-lactamases released by the lysozyme method. In addition, β-lactamase released by osmotic shock treatment was found to be unstable during storage at -20°C or during the 18 h period of iso-electric focusing at +4°C. Chloroform treatment produced similar band patterns and at least as good an enzyme yield as ultrasonic disintegration and was equally simple and fast to perform. © 1993 The British Society for Antimicrobial Chemotherapy.