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Large scale micro-and nanofabrication technologies for bioanalytical devices based on R2R imprinting

Periodic Reporting for period 3 - R2R Biofluidics (Large scale micro-and nanofabrication technologies for bioanalytical devices based on R2R imprinting)

Okres sprawozdawczy: 2018-02-01 do 2019-07-31

Roll-to-roll (R2R) technologies are mature core processes in manufacturing lines for graphical printing industry. In several other areas (e.g. electronics or optics) R2R techniques are emerging, in particular relying on recently developed roller-based nanoimprinting methods. They enable manufacturing of highly cost-effective and large-scale sheets of flexible polymer film with extremely precise structures on a micro- and nanoscale.
Areas that will benefit strongly from adopting such R2R imprinting technologies are microfluidics, biosensors, and lab-on-chip products for point of care diagnostics, drug discovery and food control. Here R2R fabrication will greatly reduce production costs and increase manufacturing capacity with respect to currently used products.
Within the European Horizon 2020 project R2R Biofluidics, a pilot production line and complete process chain for manufacturing of lab-on-chip devices is set up, based on high-throughput R2R imprinting in combination with R2R biomolecule printing and further backend process technologies.

The capabilities of these R2R production technologies is demonstrated based on two applications:
• Demonstrator 1: In-vitro diagnostic chip with imprinted microfluidic channels based on optical chemiluminescence detection using photodetectors; containing imprinted optical microstructures for light coupling and thus improving device performance.
• Demonstrator 2: Neuron-culture plates containing imprinted cavities and channels for controlled culturing and fluorescence imaging of neurons, to be applied in high throughput drug screening.

Each demonstrator development includes simulation, layout generation, mastering, shim fabrication, roll-to-roll imprinting, roll-to-roll biomolecule printing as well as inlet cutting, bonding and chip/device cutting. Both demonstrators are finished by testing of improved device performance as well as by fluidic experiments.
In the first half of the project, all developments focused on R2R imprinting of microfluidic patterns on foil substrates – the first step for manufacturing of foil based microfluidic devices. Imprint stamps of the structures for the two demonstrators were developed, imprint materials were adapted to the requirements:

Requirements of Demonstrator 1:
• Fluidics: The chip works with capillary driven liquid flow. Therefore the wetting contact angle of the used material were optimised to a water contact angle of 45° and below.
• Biofunctionalisation: Printed DNA and protein arrays.

Requirements of Demonstrator 2:
• Fluidics: Network of cell growth cavities which are connected via micro-channels. During manufacturing, the cavities need to be selectively filled with a cell adhesive solution, which then dries out and immobilizes on the channel/cavity surfaces.
• Biofunctionalisation: Cell adhesive coating, which can be selectively immobilized in the imprinted fluidic cavities.

The second half of the project focused on set-up of backend process equipment. With the newly entering partner Scienion (SCI) a microarray spotter was integrated in a R2R based manufacturing platform for microfluidic devices for the first time. BIF set-up lamination processes for manufacturing of DEM2 microtiter plates. TEC and JR set-up lamination processes on large area polymer sheet format (minimum 200 mm x 300 mm) for DEM1. BIF and JR set-up laser cutting processes for inlet and chip/device cutting.

Overview of device validation results:

GSB validated DEM1 chips by chemoluminescence measurements.
- Imprinted structures on the bottom side of the chip reduce total internal reflections and therefore allows a higher percentage of the chemoluminescence signal to leave the chip towards the detector. An increase of signal intensity by factor 1,8 was achieved.
- Imprinted reflector structures on chip top side increased the light signal which is directed towards the detector below the chip. An increase of signal intensity by factor 1,8 was achieved.
- Fully foil based chips from R2R imprinting and microarrayspotting achieved comparable results like standard chips from injection molding.

IPT validated DEM2 devices by cell cultivation.
- Selective cell growth within the imprinted cavities was successfully achieved on the materials mr-UVCur26SF (MRT) as well as JR6.1 (JR).

Overview of Exploitation and Dissemination
GSB will exploit the results of DEM1 in products with higher sensitivity as well as with full foil based design. IPT implements the micrstructured cell culture substrates in their product series. The results have been actively communicated in conferences of the microfluidics, medical sensor and cell screening community.
R2R Biofluidics enables a severe change within the microfluidic industry with the establishment of R2R manufacturing as an industrially viable fabrication technology. Within the project, the worldwide first R2R microarray spotter line for foil based microfluidics was established – which was an essential step on the way to a full implementation of R2R manufacturing of microfluidic devices.

R2R manufacturing will easily allow for throughputs > 5.000 chips/h and hence could lead to reduction of production costs by a factor of 5-10. R2R Biofluidics worked towards integrated production concepts for R2R manufacturing lines combining all relevant processing steps (imprinting, printing, lamination etc.) for microfluidic, bioanalytical consumables based on plastic foils. This is far beyond previous approaches that usually comprise just single R2R patterning steps.

Regarding the design and functionality of selected demonstrator applications
themselves, a number of innovations were realized in the project:
Novel optical outcoupling structures were integrated into a microfluidic chip for
chemiluminescence based detection of that significantly improve optical detection efficiency. A novel method for patterned functionalization of microchannels by selective wetting contrast has been established. A new class of directly biofunctionalizable R2R UV-imprinting resist was developed, and the integration of actuators (pumps/valves) to microtiterplates has been shown, enabling cell culture device with higher degree of automation (autonomous, controlled perfusion and buffer exchange) without requiring complete robotic lab automation systems. All those advancements will further contribute to significant future impact on society and public health.
For example, bacteria that are resistant to any class of antibiotics are a very real and close threat. R2R fabricated diagnostic chips will allow to rapidly screen for such pathogens in patients entering a hospital and thus allow to isolate infected or colonized patients. Such technology will prevent superbugs from spreading throughout hospitals causing catastrophic outbreaks. Such outbreaks not only cause many deaths but also a great amount of costs that public health systems have to bear.
As another example, the project will provide a device that would allow more reliable and robust cell based assays for drug screening. Using such devices, will speed up the drug production pipeline and accelerate introduction to the market. This will have a positive influence in the availability of new promising treatments for patients.
Header of R2R Biofluidic Project
Schematic of R2R imprinting of microfluidic structures
Process scheme of R2R Biofluidics - from the idea to point of care diagnostic