Multiplexed Sepsis Immunosorbent Assay Based on Flower-like CoFe₂O₄/MoS₂ with Dual Substrate Affinity

Sepsis diagnosis and prognosis are hindered by the limitations of traditional single-parameter assays, which are costly, slow, and impractical for resource-limited settings. To overcome this, we developed a novel combined nanozyme-linked immunosorbent assay (NLISA) kit for the multiplexed detection of three critical sepsis biomarkers: C-reactive protein, serum amyloid A, and D-dimer. Our approach leverages a unique flower-like CoFe₂O₄/MoS₂ nanosheet with dual substrate affinity, which serves as a highly stable and efficient nanozyme label. This sandwich-type immunocomplex enabled us to achieve robust sensitivity, specificity, and reproducibility at clinically relevant concentrations. Clinical validation using plasma samples from sepsis patients demonstrated strong concordance with the collective data from three separate enzyme-linked immunosorbent assay tests, confirming a significant improvement in the diagnostic reliability. Our NLISA kit not only simplifies multibiomarker detection but also provides a more comprehensive and accessible tool for sepsis diagnosis and monitoring, offering a powerful alternative to conventional methods.