A robust serological test is required to detect neutralizing antibodies (NAbs) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). A nanomaterial-coupled metasurface plasmon resonance (MetaSPR) biosensor is developed for rapid multiplexing quantitative tests of the half maximal inhibitory concentration (IC50) value of SARS-CoV-2 NAbs in human sera. This biosensor is applied to a MetaSPR competitive or surrogate virus neutralization test (MSPR VNT) used for quantifying NAb-mediated blockage of the interaction between the human angiotensin-converting enzyme 2 (hACE2) receptor and the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein. Novel nanomaterial, gold platinum nanoflowers (Au@Pt NFs), is combined with the optimized MetaSPR chip for dual signal amplification. The clinical utility of this biosensor is validated in 40 participants, and the results are confirmed with the IC50 values calculated using the pseudovirus neutralization test. Similarly, the neutralization titer of Omicron and Delta variants and the ability of NAbs to resist these variants are monitored. The MSPR VNT miniaturized optical biosensor instrument has a high throughput and enables one-step, rapid, and accurate quantification of SARS-CoV-2 variants or viruses causing other infectious diseases, which is applicable in vaccine trials, immune response studies, and epidemiological studies