Ultrasensitive Electrochemical Vertical Flow Immunoassay for Rapid and Simultaneous Detection of Interleukin-6 and Procalcitonin

Rapid and multiplexed detection of biomarkers plays an indispensable role in disease diagnosis. Although paper-based lateral flow immunoassays have been widely used in this field, the speediness and throughput are still challenging issues. Herein, an electrochemical vertical flow immunoassay device (eVFID) is fabricated for rapid, ultrasensitive, and multiplexed detection of inflammatory biomarkers. Working electrodes with excellent electrochemical performance and permeability properties were directly fabricated on the nitrocellulose membrane to enable both the vertical flow of the sample solution and electrochemical detection. This vertical configuration can remarkably improve the speediness of the immunoassay and effectively inhibit the cross-talk reactions among immunomolecules, thus allowing rapid and simultaneous detection of multiplexed biomarkers. Furthermore, a signal amplification strategy based on horseradish peroxidase and tetramethylbenzidine was integrated into the eVFID to substantially increase the sensitivity of the electrochemical detection. A low limit of detection of 0.1 and 0.22 pg mL^-1 was obtained for two low-abundance inflammatory biomarkers, interleukin-6 (IL-6) and procalcitonin (PCT), respectively. Finally, using a two-channel eVFID, simultaneous detection of IL-6 and PCT in human plasma samples was successfully realized within 5 min. We believe that the eVFID holds great promise for speedy and high-throughput biomarker detection at the point of care.