A one-pot proximity-triggered hybridization chain reaction ultrasensitive diagnosis platform for...

Serological detection of viral antigens and antibodies remains central to transfusion-transmitted infection (TTI) screening, but current limitations in sensitivity and operational ease demand more advanced solutions. Here we present HCR Pro, a one-pot, wash-free platform that converts target-dependent proximity binding into an enzyme-free nucleic acid amplification signal. Specifically, dual-probe recognition brings oligonucleotide-conjugated capture probes into spatial proximity, triggering initiator strand release through competitive displacement within a thermodynamically stabilized blocker complex. This initiator strand triggers a hybridization chain reaction (HCR), generating fluorescence proportional to the target concentration. Using spiked samples, HCR Pro achieves limits of detection of 10.22 fg/mL for HBsAg, 0.23 fg/mL for HCV-core Ag, and 1.38 fg/mL for HIV-1 P24 Ag and performs antibody quantification of HIV-1 Ab down to 51.75 pg/mL over a range of 4.5 × 10² to 4.5 × 10⁹ pg/mL. For a clinical sample set of 158 serum samples, HCR Pro distinguishes an infected set of samples from negative samples with an AUC value between 0.95 and 1.00, >94% sensitivity, and 100% specificity, requiring only 90 min and 15 μL of sample. The modular probe architecture allows target switching without re-optimizing the amplification backbone, positioning HCR Pro as a practical and adaptable screening platform for improving transfusion safety.