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

BACKGROUND: β-lactam resistance in Gram-negative bacteria is a significant clinical problem in the community, long-term care facilities, and hospitals. In these organisms, β-lactam resistance most commonly results from the production of β-lactamases. In Gram-negative bacilli, TEM-, SHV-, and CTX-M-type β-lactamases predominate. Therefore, new and accurate detection methods for these β-lactamase producing isolates are needed. RESULTS: E. coli DH10B cells producing SHV-1 β-lactamase and a clinical isolate of K. pneumoniae producing SHV-5 β-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 β-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.