Laser-printing of toner-based 96-microzone plates for immunoassays

Abstract

This work describes the quick and simple fabrication of toner-based 96-microzone plates by a direct-printing technology. The printer deposits a toner layer (ca. 5 μm thick) on the polyester surface which acts as a hydrophobic barrier to confine small volumes of sample on test zones (wells). A 96-microzone toner plate was explored to demonstrate its capability of performing enzyme-linked immunosorbent assay (ELISA). The detection of anti-immunoglobulin G (anti-IgG) and immunoglobulin M (IgM) antibodies has been successfully achieved in cell culture and serum samples, respectively. The use of a conventional microplate reader has allowed obtaining a limit of detection of 13 fmol of mouse IgG per zone on printed microplates. The IgM antibody has been detected in a serum sample collected from a patient infected with dengue virus. The detection of a primary infection has been provided by a microplate reader and also by a cell phone camera. Besides the bioanalytical feasibility, toner-based zones have shown good repeatability for inter-zone and intra-plate comparisons. The relative standard deviation (RSD) values for inter-zone (n = 12) and intra-plate (n = 3) comparisons were lower than 6% and 11%, respectively. Furthermore, it was found that the lifetime of each printed microplate depends on the storage temperature. The shelf life for devices stored at 10 °C has been estimated to be ca. four weeks.

Colorimetric Detection of Glucose in Biological Fluids Using Toner-Based Microzone Plates

“…5 µm (equivalent to the toner layer thickness). 32 When compared to wax printing technology, 35 recognized as one of the simplest methods to produce paper-based analytical devices, the laser printing allows the fabrication of microzone plates in a single printing step using simpler instrumentation. In this current report, zones were filled with a cellulose paste to ensure the retention of the added reagents and to provide a better color distribution on the whole zone.…”

“…Moreover, this strategy improved the repeatability of colorimetric measurements through digital images captured with a scanner and prevented the interference of external light commonly observed in the drop formed inside microzones. 32 Glucose colorimetric assay was then carried out using toner-based microzones filled with cellulose paste. The reaction is based on glucose conversion to gluconic acid and hydrogen peroxide by glucose oxidase (GOx) enzyme in the presence of HRP, which catalyzes the reduction of hydrogen peroxide and the oxidation of iodide to molecular iodine.…”

“…32 Microplates were drawn in Corel Draw software version 13.0 and then printed on a polyester film surface using a laser printer with 1200-dpi resolution (model LaserJet 1102w, Hewlett-Packard). The toner provided a hydrophobic barrier restricting the area for bioassays.…”

“…This platform has been successfully used for capillary-driven flow based clinical assays 31 and dengue diagnostics through enzyme-linked immunosorbent assay (ELISA) on printed zones. 32 When compared to capillary-driven toner-based microfluidic devices, the laser printing of microzone array on transparency films is based on a single printing step, 32 while the fabrication of multilayer toner-based microfluidic devices require the laser cut of an intermediary transparency film, an alignment step and, finally, a thermal lamination to promote the channel sealing. 31 In this current report, we describe the use of tonerbased microzone plates for glucose analysis in biological fluids using colorimetric measurements.…”

Polymeric-Based In Vitro Diagnostic Devices

“…First, Oliveira et al [108] described the quick and simple fabrication of toner-based 96-microzone plates to detect dengue virus. They created detection zones (wells) by printing a hydrophobic toner layer (ca.…”