Background: Enzyme-linked immunosorbent assays (ELISA) are considered the gold standard in the demonstration of various immunological reactions with an application in the detection of infectious diseases such as during outbreaks or in patient care.
Objective: This study aimed to produce an ELISA-based diagnostic with an increased sensitivity of detection compared to the standard 96-well method in the immunologic diagnosis of infectious diseases.
Methods: A '3DStack' was developed using readily available, low cost fabrication technologies namely nanoimprinting and press stamping with an increased surface area of 4 to 6 times more compared to 96-well plates. This was achieved by stacking multiple nanoimprinted polymer sheets. The flow of analytes between the sheets was enhanced by rotating the 3DStack and confirmed by Finite-Element (FE) simulation. An Immunoglobulin G (IgG) ELISA for the detection of antibodies in human serum raised against Rubella virus was performed for validation.
Results: An improved sensitivity of up to 1.9 folds higher was observed using the 3DStack compared to the standard method.
Conclusions: The increased surface area of the 3DStack developed using nanoimprinting and press stamping technologies, and the flow pattern between sheets generated by rotating the 3DStack were potential contributors to a more sensitive ELISA-based diagnostic device.
Keywords: Enzyme-linked Immunosorbent Assay (ELISA); High surface-to-volume ratio diagnostics; microfluidics; nano-imprinting; press-stamping; surface plasma treatment.