Rapid method of extraction and analysis of extended-spectrum β-lactamases from clinical strains of Klebsiella pneumoniae

The extraction of periplasmic β-lactamases from Gram-negative bacilli is a necessary preliminary step to analytical isoelectric focusing. Previously described methods are time-consuming and require large amounts of broth. We describe a lysozyme-based method which needs just 5 mL broth and requires less than 24 h to perform. The method was reproducible in extracting β-lactamases from reference strains containing known β-lactamases. We applied the method to a collection of more than 70 extended-spectrum β-lactamase-producing isolates from a multinational study of bacteremic isolates of Klebsiella pneumoniae. Further studies are being undertaken to assess the method's applicability to other bacterial species.