The advancement of sensing systems and methodologies that have the ability to provide continuous results in real time, at low cost and high throughput, has been driven by the increasing demand for cell-based assays to figure out the interaction that takes place between cells and biological and chemical compounds or materials. These requirements can be satisfied by using a combination of biological components, such as proteins, with electrically conductive interdigital structures, which is a non-invasive methodology.
Sensors for use in biological applications that have interdigital structures already prevail at present. Indeed, currently, there exist a number of commercial variants that contain electrically conductive structures formed of biocompatible metals (for example, platinum or gold) and are usually produced through photolithographic techniques. Since viable cells are examined by using cell-based sensors, a biocompatible interface to a fluidic system is made necessary, and can essentially be achieved by bonding the wells to the printed circuit board containing the electrically conductive interdigital structures. However, it must be noted that these structures are usually very costly to make due to the complicated production technique of the biocompatible interdigital structures.
The BIOGRAPHY Project
The BIOGRAPHY research project has led to the development of innovative production methods using Haydale functionalized graphene ink for a quick transfer of research results from materials science to industrial applications. At present, initial investigations of this project are ongoing in relation to the usability of the printed biosensors in the proposed fields of application.
Cost-efficient manufacture of the sensors is enabled by the application of a roll-to-roll gravure printing technology, which involves bonding printed foils with bottomless well plates to arrange the sensors in a well-plate format, as illustrated in Figure 1.
Figure 1. 24 Printed interdigital electrodes.
These sensors are developed through the sequential printing of two substances, which involves printing the interdigital structures with the help of conductive ink that contains graphene platelets. The second step in the process involves printing an additional protein layer onto the electrode structures to enhance the adhesion of cells to the sensor, for instance, as a confluent monolayer. Register control, which is otherwise believed to be the synchronization of the printing units with one another, is adopted to make sure that the layers are precisely aligned with each other, which is specifically significant for ensuring the quality and function of the sensors.
Haydale’s Graphene-Containing Ink
Recently, Haydale has developed an innovative graphene-containing ink for gravure printing and investigated its suitability, conductivity, cytotoxicity, and biocompatibility. It was discovered that the ink successfully satisfied the demands pertaining to printability and application in a cell-based sensor. This innovative ink can be used to develop printed structures with a surface resistance of 10 Ω/sq based on a layer thickness of 25 μm. The print cylinder, which has the ability to produce structures with lateral dimensions less than 10 μm, was created by using a newly developed micro-engraving machine with an ultra-short pulse laser.
For producing the sensors, a compact roll-to-roll printing system with standard printing parameters for two-color printing has been assembled. Furthermore, an integrated corona unit to carry out surface activation of the substrate is fitted in the system, and a near-infrared drying unit is provided in each of the two printing units. The graphene ink, the plant, and the developed processes are all appropriate for high-speed printing of conductive graphene electrodes, designed as interdigital structures. This technology can be used to print even very fine electrode structures that have a width of 52 µm and a distance of 52 µm between the adjacent fingers.
The graphene-containing ink is electrically conductive, nano-enhanced, and biocompatible as a printed layer. Cytotoxicity tests conducted according to ISO 10993-5 with varied cell lines demonstrated the biocompatibility of the ink, which resulted in the modification of the ink to enable its usage through rotary gravure printing. In order to make sure the definition and accuracy of the printing technology are not affected at high speed, the ink had to be prepared such that it dries quickly and has an appropriate viscosity.
Composition of the Graphene-Containing Ink
The main constituents of the ink are chief component types of a resin (or binder), solvents, and additives, including pigments or other functional additives. The blend of targeted characteristics was achieved by carefully choosing the constituents, particularly the addition of Haydale’s Graphene Nanoplatelet (GNP) material as the main active ingredient. GNPs are carbon nanomaterials with high aspect ratio and are processed and surface-functionalized by using Haydale’s patented HDPlas® plasma-based technology.
The low-energy HDPlas® process introduces a functional chemistry in a distinctively benign way that minimizes the amount of damage to the nanomaterial structure, thus enhancing the potential to confer the nanomaterial characteristics to the final ink. This chemical functionalization process enhances the efficiency of the diffusion into the carrier resin used in the ink since this process enables the ink to more readily present the preferred properties mentioned earlier. The ink was produced by adopting a range of mixing and dispersion techniques, which involved mixing GNPs into the resin vehicle and then subjecting them to a sequence of compounding processes to guarantee sufficient uniformity through the ink.
The ensuing biocompatible ink has a paste-like viscosity that can be diluted with a suitable solvent. A number of factors have a direct impact on the choice of the solvent, for example, printing speed, chemical compatibility, and drying factors, such as time and temperature, to name a few. At an appropriate solvent concentration, the viscosity of the ink was 0.1 Pa*s; however, the surface tension was 31–36 mN/m and the square resistance was established for prints made with the new Haydale graphene ink and a two-color printing machine. The well depth of the test cylinder was 60 μm. The thickness of the printed layer was approximately 5.5 μm and its measured square resistance was about 100 Ω/sq. Normalization to a layer thickness of 25 μm led to a square resistance of around 20 Ω/sq.
As illustrated in Figure 2, electrically conductive interdigital structures of differing geometries were successfully printed. It has to be noted that even the smallest electrode structures that have widths of 50 µm and a 50 µm distance between adjacent electrodes could be produced through gravure printing.
Figure 2. Printed interdigital electrodes. Electrode width and gap: 50 µm.
The sensors developed as part of the BIOGRAPHY project can be applied for toxicity studies and also for validating the effectiveness of anti-infective agents, such as antiviral substances. In such cases, a confluent cell monolayer is formed on the interdigital electrode-based sensor by using the indicator cell lines and is electrically insulated. When the electrically insulating, confluent cell layer is destroyed by the added substances, it will thus show, for instance, the cytotoxicity of these substances. Visualization of the reduction of the electrical resistance of the interdigital structure helps identify this cellular response.
In the case of the second potential application, viruses, and not toxic substances, are added to the electrically insulating cell layer, thus resulting in a morphological modification of the cells or a detachment of the cells from the cell monolayer, which is otherwise known as the cytopathic effect (CPE). When antiviral substances are added, the viral replication in the cells or the infection of the cells is inhibited, thereby inhibiting the CPE. This scenario can be helpful in deriving the changes in the measured impedance values from the achieved inhibition of the CPE and hence the efficacy of the antiviral substances.
Innovate UK, the UK’s innovation agency, has funded the BIOGRAPHY project. Innovate UK primarily works with companies, people, and partner organizations to discover and drive the scientific and technological innovations that will boost the growth of the UK economy. The project has also been funded by the German Federal Ministry of Education and Research (BMBF) and managed by the Project Management Agency Karlsruhe (PTKA).
This information has been sourced, reviewed and adapted from materials provided by Haydale Limited.
For more information on this source, please visit Haydale Limited.