Gene mutation is associated with cancer and the dissemination of circulating biomarkers through the blood represents a vital transitional step that exemplifies the shift from localized to systemic disease. The...

Paper-based analytical devices have been substantially developed in recent decades. Many fabrication techniques for paper-based analytical devices have been demonstrated and reported. Herein we report a relatively rapid, simple, and inexpensive method for fabricating paper-based analytical devices using parafilm hot pressing. We studied and optimized the effect of the key fabrication parameters, namely pressure, temperature, and pressing time. We discerned the optimal conditions, including pressure of 3.8 MPa (3 tons), temperature of 80oC, and 3 minutes of pressing time, with the smallest hydrophobic barrier size (821 µm) being governed by laminate mask and parafilm dispersal from pressure and heat. Physical and biochemical properties were evaluated to substantiate the paper functionality for analytical devices. Wicking speed in the fabricated paper strips was slightly slower than that of non-processed paper, resulting from reducing paper pore size. A colorimetric immunological assay was performed to demonstrate the protein binding capacity of the paper-based device after exposure to pressure and heat from the fabrication. Moreover, mixing in two-dimensional paper-based device and flowing in a three-dimensional counterpart were thoroughly investigated, demonstrating that the paper device from this fabrication process is potentially applicable as analytical devices for biomolecule detection. Fast, easy, and inexpensive parafilm hot press fabrication presents an opportunity for researchers to develop paper-based analytical devices in resource-limited environments.

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This paper reports the development of fluorescent-linked immunosorbent paper-based assay for exosome detection. The paper-based device was fabricated with sandwich lamination for easy handling and was coated with exosome-specific antibody as a biosensing platform to detect exosome sample from the cell culture media. This assay employed fluorescent detection which is followed by tagging fluorophore-conjugated detecting antibody on exosome samples. The fluorescent readout was evaluated and quantified from image processing software. This assay can detect high concentration of exosome samples (~ 1010 exosome/mL). However, this assay has encountered various challenges. First, the exosome concentration prepared from cell culture media from cancer-derived ovarian and mesothelial cell lines may be insufficient to reach detectable range. Second, chemical contamination from exosome isolation kits may affect assay sensitivity. Therefore, assay optimization and minimizing chemical contamination are required which could enhance assay specificity and sensitivity.

We report on an immunofluorescent paper-based assay for the detection of severe acute respiratory symptom coronavirus 2 (SARS-CoV-2) humanized antibody. The paper-based device was fabricated by using lamination technique for easy and optimized handling. Our approach utilises a two-step strategy that involves (i) initial coating of the paper-electrode with recombinant SARS-CoV-2 nucleocapsid antigen to capture the target SARS-CoV-2 specific antibodies, and (ii) subsequent detection of SARS-CoV-2 antibodies using fluorophore-conjugated IgG antibody. The fluorescence readout was observed with fluorescence microscopy. The images were processed and quantified using a MATLAB program. The assay can selectively detect SARS-CoV-2 humanized antibodies spiked in PBS and healthy human serum samples with the relative standard deviation of approximately 6.4% (for n = 3). It has broad dynamic ranges (1 ng to 50 ng/µL in PBS and 5 to 100 ng/µL in human serum samples) for SARS-CoV-2 humanized antibodies with the detection limits of 2 ng/µL (0.025 IU/mL) and 10 ng/µL (0.125 IU/mL) in PBS and human serum samples, respectively. We believe that our assay has the potential to be used as a simple, rapid, and inexpensive paper-based diagnostic device with a portable fluorescent reader to provide point-of-care diagnosis. This assay can be used for rapid examination of a large batch of samples toward clinical screening of SARS-CoV-2 specific antibodies as a confirmed infected active case or to evaluate the immune response to a SARS-CoV-2 vaccine.