Integration of Laser-Induced Graphene Strain Gauges and Stimulation Electrodes into 3D Muscle on a Chip Devices

The emerging technique of direct laser carbonization of polymers to convert them into conductive graphene foam has opened new possibilities for prototyping and fabrication of electronic devices. For example, polyimide (PI) undergoes a photothermal conversion into a 3-dimensional (3D) carbon foam called laser-induced graphene (LIG). This technically simple procedure can be implemented for the photolithography-free fabrication of electronic devices in the absence of a cleanroom, with faster turnover of prototypes, and at a lower cost. The technique has similar spatial resolution to inkjet printing but is more straightforward in implementation.<br/>Muscle on a Chip models have been developed to gain insight into disease mechanisms from the molecular to the tissue scale and test new therapies. Systems that engineer muscle tissue in 3D bundles are especially beneficial for inducing more physiologically relevant tissue architecture. 3D muscle bundles are commonly fabricated by injecting a mixture of muscle cells and hydrogel into an elongated chamber with two flexible anchors. The muscle cells compact into an aligned 3D bundle that mimics native muscle tissue. The flexible anchors are embedded into the tissue during compaction and deflect with muscle contractions. However, the conventionally used method to evaluate muscle contractions by optically monitoring the anchors has low throughput, and limited options for continuous monitoring. To overcome these bottlenecks in Muscle on a Chip systems, we integrated LIG electrodes in our devices. Based on the same engraving technique, we fabricated electrodes for the electrical stimulation and the contractility measurement.<br/>To fabricate LIG on PI films (LIG-PI), we used an Epilog Mini laser engraver equipped with a CO₂ 10.6 μm wavelength laser with maximum power of 30 W. By engraving patterns as an array of raster dots or overlapping vector “lines”, and varying laser power and speed, we obtained LIG layers with sheet resistance from 120±17 Ω/sq to 32±4 Ω/sq.<br/>This electrode demonstrated a charge injection capacity in the range of 0.2 mC/cm² for cathodic and 0.1 mC/cm² for anodic current pulse. Stability of the material after thousands of stimulation pulses was confirmed by measuring Ferricyanide-ferrocyanide oxidation/reduction currents in cyclic voltammetry and calculating the electrochemically active surface area.<br/>We transferred LIG foam into polydimethylsiloxane (PDMS) films by pouring liquid PDMS on LIG-PI, curing it until solid, and peeling off the PI film. This created a stretchable PDMS film embedded with LIG foam (LIG-PDMS), the conductivity of which responded to the stress applied to the layer. We demonstrated that LIG-PI and LIG-PDMS were not toxic by culturing HL-1 cardiomyocyte-like cells on the materials for 1 week.<br/>To fabricate the strain gauge, we engraved LIG in a serpentine line pattern on PI films and transferred it onto PDMS. We further shaped this layer by laser cutting excess PDMS to create flexible anchors for the 3D tissue device, which should change resistivity upon deflection caused by the contracting muscle. To fabricate muscle chambers, we replica molded top and bottom pieces of PDMS from templates 3D printed in a Digital Light Processing printer. After 7 days of culture required for tissue compaction, LIG-PI stimulation electrodes were used to induce contractions of a primary muscle bundle at 2 Hz (twitch) and 20 Hz (tetanus).<br/>We plan to use the developed device to evaluate physiological responses of two tissue models: healthy and diseased cardiomyocytes derived from human induced pluripotent stem cells, and primary skeletal muscle cells with continuous electrical stimulation and monitoring of muscle maturation.