Microfluidic paper-based multiplex colorimetric immunodevice based on the catalytic effect of Pd/Fe3O4@C peroxidase mimetics on multiple chromogenic reactions

•Paper-based colorimetric immunosensor for CEA and α-AFP based on two detection layers was developed.•The peroxidase-like catalytic performance of Pd/Fe3O4@C MNPs was used for signal amplification.•Chitosan and FLAuNPs were employed as the carriers of the primary antibodies.

In this report, a non-toxic method was proposed for the simple synthesis of palladium nanoparticles (Pd)/Fe3O4@C peroxidase mimetics by virtue of in situ growth of Pd nanoparticles on Fe3O4@C magnetic nanoparticles. And a microfluidic paper-based multiplex colorimetric immunodevice (named α-sheet) was developed by site-selectively immobilizing multiple antigens owing to its intrinsic high-efficiency catalytic activity of peroxidase mimetics to multiple chromogenic reactions. The immunosensor platform was prepared by growing a layer of flower-like gold nanoparticles which could entrap the primary antibodies onto paper sensing zones, and the as-prepared Pd/Fe3O4@C peroxidase mimetics was used to label secondary antibodies. In the presence of 3,3′,5,5′-tetramethylbenzidine and o-phenylenediamine chromogenic substrates, Pd/Fe3O4@C peroxidase mimetics catalyzed chromogenic reactions and showed different colors with respective intensity. To precisely identify the intensity, a piece of black wax printed chromatographic paper with three observing windows (named β-sheet) was flatted on α-sheet. Under the optimal condition, the proposed multiplex colorimetric immunodevice displayed wide linear ranges from 0.005 to 30 ng mL−1 with low detection limits of 1.7 pg mL−1 for carcinoembryonic antigen (CEA) and α-fetoprotein (α-AFP). Meanwhile, the proposed method provided provided a non-toxic, low-cost and promising tool for point-of-care diagnosis.

Graphical abstractPd/Fe3O4@C magnetic nanoparticles and flower-like AuNPs were prepared to immobilize Ab1 and Ab2 respectively and combined to fabricate a sandwich-type paper-based colorimetric immunosensor for multiplexed measurement of biomarkers.Figure optionsDownload full-size imageDownload as PowerPoint slide