Immuno-transcription-amplified single microbead assay for protein and exosome analysis through an S9.6 antibody-nucleic acid recognition strategy.

High-sensitive detection of circulating biomarkers is in high demand because many of them are found at low concentrations in bioliquids. Herein, we report an immuno-transcription-amplified single microbead (MB) assay (IT-SMA) based on the specific S9.6 antibody-DNA/RNA hybrid recognition strategy for the sensitive and universal quantification of protein biomarkers. This design rationally converts the immunoreaction events into amplified nucleic acid transcription to produce numerous RNA molecules, which can efficiently enrich fluorescent signals onto a single MB through a specific S9.6 antibody-DNA/RNA hybrid recognition mechanism, enabling sensitive protein analysis. This method exhibits excellent specificity and high sensitivity for protein analysis with a low detection limit at the fg/mL level. Furthermore, the S9.6 antibody-aided IT-SMA allows for universal detection of various proteins and even exosomes, testing target proteins in serum samples, and differentiating cancer patients from healthy individuals by directly analyzing the exosomes in human blood samples. These features make the IT-SMA strategy a promising tool for the quantitative detection of a variety of biomarkers toward precision diagnostics.