Functional DNA-enhanced quantum dots biosensor for rapid and sensitive β-lactamase detection in Haemophilus influenzae.

β-Lactamase (BL) detection plays a significant role in guiding treatment for bacterial infections. In this study, a BL detection method was developed by utilizing DNA-regulated cadmium telluride quantum dots (CdTe QDs) for selective recognition. Experiments demonstrated that BL hydrolyzed penicillin G (PG) to generate penicilloic acid (PA), which effectively reduced Cu2+ to Cu+. The CdTe QDs exhibited high selectivity toward both Cu2+ and Cu+. Systematic investigations revealed that different DNA configurations could specifically modulate this recognition process; notably, DNA showed selective binding with Cu2+ but not with Cu+. DNA encapsulated Cu2+, preventing its reaction with CdTe QDs while allowing only Cu+ to react with the QDs, thereby significantly reducing background signal interference. The functional DNA nanostructure, protein-scaffolded DNA (PS DNA) nanonet, exhibited superior Cu2+ loading capacity compared to other structures, enhancing the detection sensitivity for PA to the pM level. This approach achieved detection limits of 74.6 mU/L for BL and 590 CFU/mL for BL-positive Haemophilus influenzae (Hi), and the entire assay was completed in only 1.1 h. A total of 60 clinical samples were analyzed to validate this technique, demonstrating good consistency with nitrocefin tests and whole genome sequencing (WGS).