RESUMO
ABSTRACT: With the Covid-19-based global pandemic that started in the beginning of 2020, the vital importance of accelerated, reliable and affordable virus testing systems has once again become clearer. Besides, we all learned very well that the disposable biochips, to be used in these in vitro diagnostic (IVD) testing systems, supposed to be produced in large amounts in a very short time to be widely available for the use of humanity to save more and more lives. That is why; roll-to-roll (R2R) polymer structuring manners offer such large quantities for the production of in vitro biochips. Our technology, based on R2R UV nanoimprint lithography (UV-NIL), has superior features. Via our pilot line, robust 7500 biochip components per 100 meter of a flexible, polymer foil coated with a UV curable photo-resin (i.e., parts with capillary fluidic channels or optical structures for IVDs) can be generated. This study shows an example of a prototype of a R2R UV-NIL generated chip: a foil, capillary flow-based IVD biochip for multiplexed DNA detection purposes (i.e., a Lab-on-a-Foil device). The biochip performance was further increased dramatically by integrating UV-NIL produced retro-reflective microstructures, which reflects the light back, to its design to enhance optical signal detection in a commercial IVD device, detecting DNA on a chemiluminescent-reaction basis.
RESUMO
Roll-to-roll UV nanoimprint lithography has superior advantages for high-throughput manufacturing of micro- or nano-structures on flexible polymer foils with various geometries and configurations. Our pilot line provides large-scale structure imprinting for cost-effective polymer biochips (4500 biochips/hour), enabling rapid and multiplexed detections. A complete high-volume process chain of the technology for producing structures like µ-sized, triangular optical out-couplers or capillary channels (width: from 1 µm to 2 mm, height: from 200 nm up to 100 µm) to obtain biochips (width: 25 mm, length: 75 mm, height: 100 µm to 1.5 mm) was described. The imprinting process was performed with custom-developed resins on polymer foils with resin thicknesses ranging between 125-190 µm. The produced chips were tested in a commercial point-of-care diagnostic system for multiplexed DNA analysis of methicillin resistant Staphylococcus aureus (e.g., mecA, mecC gene detections). Specific target DNA capturing was based on hybridisation between surface bound DNA probes and biotinylated targets from the sample. The immobilised biotinylated targets subsequently bind streptavidin-horseradish peroxidase conjugates, which in turn generate light upon incubation with a chemiluminescent substrate. To enhance the light out-coupling thus to improve the system performance, optical structures were integrated into the design. The limits-of-detection of mecA (25 bp) for chips with and without structures were calculated as 0.06 and 0.07 µM, respectively. Further, foil-based chips with fluidic channels were DNA functionalised in our roll-to-roll micro-array spotter following the imprinting. This straightforward approach of sequential imprinting and multiplexed DNA functionalisation on a single foil was also realised for the first time. The corresponding foil-based chips were able to detect mecA gene DNA sequences down to a 0.25 µM concentration.