Dual-signal ELISA-SERS immunosensor based on a multifunctional magnetic nanozyme for sensitive detection of microcystin-LR

Microcystin-LR (MC-LR) is a potent cyanotoxin that threatens environmental safety and public health. Here, we developed a dual-signal ELISA-SERS immunosensor based on a multifunctional magnetic nanozyme, MSNP@Ag/Pt-4-MBN (MSAT), for highly sensitive MC-LR detection. The probe integrates magnetic mesoporous silica, Ag/Pt bimetallic nanostructures, and 4-mercaptobenzonitrile (4-MBN), with Pt nanoparticles driving TMB/H₂O₂ colorimetry and Ag nanostructures enhancing the SERS signal of the embedded 4-MBN reporter. After conjugation with the A2.3-SBP nanobody, the probe was applied in a competitive immunoassay. The colorimetric signal was first measured, followed by magnetic washing and subsequent SERS measurement. Both absorbance at 450 nm and SERS intensity at 1076 cm⁻¹ decreased with increasing MC-LR concentration over a broad range of 0.01–1000 μg/L. The limits of detection were 0.00455 μg/L for colorimetry and 0.00541 μg/L for SERS. The immunosensor showed excellent selectivity, repeatability, and inter-batch reproducibility, together with satisfactory recoveries and good agreement with HPLC. By integrating magnetic separation, catalytic amplification, and dual-readout cross-validation, this platform provides reliable MC-LR monitoring in food and environmental samples.