Evaluation of Viral Collection Efficiency with Antibody-Modified Magnetic Particles by Polymerase Chain Reaction...

Polymerase chain reaction (PCR) is the primary method for virus detection; however, its complex preprocessing has prompted research into simpler immunoassay-based approaches. Among these, techniques using antibody-modified magnetic particles, exemplified by digital ELISA, provide ultra-high sensitivity comparable to PCR by efficiently capturing trace viruses and enabling concentration, washing, and transfer to microreactors. In this study, we evaluated the virus capture efficiency of antibody-modified magnetic particles based on quantitative PCR (qPCR). Influenza A virus (H1N1/A/Puerto Rico/8/1934) was tested with 1 μm magnetic beads modified with HA1 antibodies. As quantification becomes unreliable and difficult in an extremely low-concentration range near the detection limit of qPCR, low-concentration viral suspensions (10⁵ copies/mL) were mixed with particle dispersions (up to 5 × 10⁸ particles/mL) for 10 min, followed by magnetic separation and washing, and the remaining virus in each fraction was analyzed by qPCR. At the highest particle concentration, capture rates exceeded 80% relative to the initial suspension, indicating near-complete capturing when considering free nucleic acids. Time-course analysis showed that the capture rate reached saturation within 2 min, with approximately 90% of the saturation at 1 min. Furthermore, kinetic modeling of magnetic bead-virus binding reproduced experimental data. These findings demonstrate that short mixing times with high particle concentrations enable efficient virus capture, contributing to the development of rapid and highly sensitive immunoassay systems.