A modular and universal platform for neutralizing-antibody profiling by quantifying antigen-receptor interactions via qPCR.

Rapid and accurate detection of neutralizing antibody (nAb) is essential for assessing vaccine efficacy and immune protection against infectious diseases. In the present study, we developed a neutralization-antibody detection qPCR platform (NAD-qPCR) that quantifies nAb potency by integrating antigen-receptor binding specificity with qPCR sensitivity. Using the SARS-CoV-2 nAb as a proof-of-concept target, this system comprised a hybrid probe with the viral RBD as an antigen module covalently conjugated to a reporter DNA, and magnetic beads functionalized with the ACE2 mimic mini-protein as a receptor module, thereby recapitulating the natural antigen-receptor interactions. With this design, the neutralizing capability of nAb against the antigen was transformed into a competitive reduction of the probe binding and DNA amplification signal by qPCR. The NAD-qPCR demonstrated robust performance in quantifying commercial SARS-CoV-2 nAb with a LOD of 4 ng/μL. Furthermore, it was demonstrated to be capable of effectively profiling a large number of vaccinated donor serum with broad neutralization activity. Notably, the hybrid probe and the capture beads were designed modularly, which allows for straightforward adaptation to other pathogens by replacing the module component. This platform offers a rapid, high-throughput, and universally applicable approach for nAb detection in vaccine development and immune monitoring.