Microtrap-assisted microfluidic magnetic separation and concentration for ultrasensitive immunoassays of biomarkers

Precise and accurate quantitation of important biomarkers is significant, especially in early-stage diseases diagnosis. To realized effective biosample preparation and trace-level biomarker detection, a microtrap-assisted microfluidic magnetic immunoassays (μMI) method was developed in this work. A microtrap was fabricated inside the straight microchannel of μMI device to help magnetic separation and concentration of immunocomplexes. These immunocomplexes were enriched in microtrap of μMI device to accomplish selective and sensitive biomarker detection. Horseradish peroxidase-labeled magnetic beads were employed to evaluate assay feasibility and microtrap effect on assay sensitivity. The microtrap-assisted μMI was then applied for model biomarkers detection. The limits of detection of μMI were 0.025 pg/mL for monocyte chemoattractant protein-1 (MCP-1) and 0.021 pg/mL for matrix metalloproteinase-9 (MMP-9), which corresponded up to 2014-fold sensitivity improvement compared to their standard microwell enzyme-linked immunosorbent assay (ELISA) results. In addition, the selectivity and reproducibility of microtrap-assisted μMI were confirmed. In clinical serum sample analysis, recoveries of 91.3%-106.7% with relative standard deviations less than 6.1% were obtained for MCP-1 and MMP-9, and method accuracy was verified by commercial ELISA kit. The developed μMI can accomplish ultratrace biomarker detection offering practical tool for laboratorial and clinical research.