A modular β-lactamase-based thermistor biosensor platform for rapid plasma monitoring of major β-lactam antibiotic classes.

Biosensors represent a promising approach for rapid therapeutic drug monitoring (TDM) of β-lactam antibiotics. However, no single biosensor strategy has yet been established to cover all four major β-lactam classes: penicillins, cephalosporins, carbapenems, and monobactams. Recently, an enzyme thermistor biosensor using New Delhi metallo-β-lactamase-1 (NDM-1) as the biorecognition element enabled the detection of penicillins, cephalosporins, and carbapenems in spiked plasma and supported accurate pharmacokinetic profiling of cefuroxime in treated patients. Here, we developed a CTX-M-14-coupled enzyme thermistor biosensor and compared its performance with that of the NDM-1 biosensor for detecting two penicillins, four extended-spectrum oxyimino-cephalosporins, one carbapenem, and one monobactam. Both β-lactamase-coupled biosensors achieved rapid response times of approximately 5 min. Across a plasma concentration range of 6.25-200 mg/L, the CTX-M-14-coupled biosensor quantified all tested penicillins, oxyimino-cephalosporins, the carbapenem, and the monobactam. In contrast, the NDM-1-coupled biosensor quantified penicillins, oxyimino-cephalosporins, and carbapenems, but not the monobactam. Although CTX-M-14 generally produced lower response signals than NDM-1 for most oxyimino-cephalosporins and the carbapenem, matrix effects for penicillins, oxyimino-cephalosporins and the monobactam were comparable between the two biosensors, with a significant difference observed only for the carbapenem. Plasma cefotaxime concentrations measured by the CTX-M-14-coupled biosensor in four treated patients closely matched UPLC-MS/MS results. Collectively, these findings support the CTX-M-14-coupled thermistor biosensor as a rapid and accurate approach for quantifying penicillins, oxyimino-cephalosporins, carbapenems, and monobactams in plasma, and highlight enzyme thermistor biosensing as a flexible platform for TDM of full-spectrum β-lactam antibiotics.