Detection of SHV β-lactamases in Gram-negative bacilli using fluorescein-labeled antibodies

Background: beta-lactam resistance in Gram-negative bacteria is a significant clinical problem in the community, long-term care facilities, and hospitals. In these organisms, beta-lactam resistance most commonly results from the production of beta-lactamases. In Gram-negative bacilli, TEM-, SHV-, and CTX-M-type beta-lactamases predominate. Therefore, new and accurate detection methods for these beta-lactamase producing isolates are needed. Results: E. coli DH10B cells producing SHV-1 beta-lactamase and a clinical isolate of K. pneumoniae producing SHV- 5 beta-lactamase were rendered membrane permeable, fixed and adhered to poly-L-lysine coated slides, and stained with purified polyclonal anti-SHV antibodies that were fluorescein labeled. E. coli DH10B cells without a bla(SHV) gene were used as a negative control. The procedure generated a fluorescence signal from those slides containing cells expressing SHV beta-lactamase that was sufficient for direct imaging. Conclusion: We developed a rapid and accurate method of visualizing the SHV family of enzymes in clinical samples containing Gram-negative bacilli using a fluorescein-labeled polyclonal antibody.