Cost-Effective and Handmade Paper-Based Immunosensing Device for Electrochemical Detection of Influenza Virus

Although many studies concerning the detection of influenza virus have been published, a paper-based, label-free electrochemical immunosensor has never been reported. Here, we present a cost-effective, handmade paper-based immunosensor for label-free electrochemical detection of influenza virus H1N1. This immunosensor was prepared by modifying paper with a spray of hydrophobic silica nanoparticles, and using stencil-printed electrodes. We used a glass vaporizer to spray the hydrophobic silica nanoparticles onto the paper, rendering it super-hydrophobic. The super-hydrophobicity, which is essential for this paper-based biosensor, was achieved via 30-40 spray coatings, corresponding to a 0.39-0.41 mg cm^-2 coating of nanoparticles on the paper and yielding a water contact angle of 150° ± 1°. Stencil-printed carbon electrodes modified with single-walled carbon nanotubes and chitosan were employed to increase the sensitivity of the sensor, and the antibodies were immobilized via glutaraldehyde cross-linking. Differential pulse voltammetry was used to assess the sensitivity of the sensors at various virus concentrations, ranging from 10 to 10⁴ PFU mL^-1, and the selectivity was assessed against MS2 bacteriophages and the influenza B viruses. These immunosensors showed good linear behaviors, improved detection times (30 min), and selectivity for the H1N1 virus with a limit of detection of 113 PFU mL^-1, which is sufficiently sensitive for rapid on-site diagnosis. The simple and inexpensive methodologies developed in this study have great potential to be used for the development of a low-cost and disposable immunosensor for detection of pathogenic microorganisms, especially in developing countries.