Real-Time In Vitro Monitoring of Cell Populations

In the ongoing Young Investigator project (YIP) funded by the Villum Foundation, researchers in the Biomaterial Microsystems group are developing smart microwells and microfluidic systems that will make it possible to monitor the behaviour of cell populations directly while they are being cultured. This is achieved by integrating carbon microelectrodes at the bottom of the microwells.

These microelectrodes serve as electrochemical sensors. Adhesion and growth of cells on the electrodes result in a change of the electrical resistance of the system which can be recorded using so-called electrochemical impedance spectroscopy (EIS). Additionally, the release of electrochemically active substances from the cells either spontaneously or after addition of external stimuli such as drugs or toxins can be measured. 

Pyrolytic 3D carbon microelectrodes

Associate Professor Stephan Sylvest Keller, head of the Biomaterial Microsystems group says: “We have chosen to work with carbon since it is an excellent material for electrochemistry – it is chemically inert and highly biocompatible. We make the carbon electrodes by so-called pyrolysis where we convert a polymer precursor template into pyrolytic carbon by heating it to high temperature – more than 900 °C - in nitrogen atmosphere”.

The final shape and properties of the carbon microelectrode are determined by the geometry and material properties of the polymer template and the pyrolysis conditions. This allows the fabrication of a large variety of carbon microelectrodes. As a particular advantage, it is possible to fabricate carbon microelectrodes with a 3D structure (see figure 2) by pyrolysis if the precursor template is a 3D polymer microstructure. In their natural environment, cells prefer to grow and interact in a 3D environment but observation of cells in a 3D culture with traditional methods is not possible. 3D carbon electrodes have the potential to serve as 3D cell scaffolds and 3D electrochemical sensors for cell monitoring at the same time. 

Figure 2: 3D carbon microelectrodes with 50 µm high pillars supporting a 5 µm thick grid with lattice of 50 µm

Carbon - the new black for in vitro cell models?
Integration of microelectrodes in microwells makes it possible for the researchers to monitor cell adhesion, growth and apoptosis (cell death). Additionally, with electrochemical methods it is possible to directly detect molecules released by the cells such as mediators for cell-to-cell communication.

With a carefully tailored electrode design, it is thus possible to measure the growth and interactions of cells directly while and where it happens. This can be useful in a number of areas. Stephan Sylvest Keller says that “At the moment, a part of the activities in the research group focuses on in vitro monitoring of bone cells to potentially accelerate the development of drugs for the treatment of osteoporosis. Here 3D pyrolytic carbon microelectrodes are interesting because their mechanical properties and geometry closely resemble osteoporotic bone tissue”.

Another possible application is the monitoring of human neural stem cells and their ability to release dopamine. This is relevant for the development of methods for the treatment of Parkinson’s disease and it is currently being explored in collaboration with the Bioanalytics research group at DTU Nanotech.

The researchers are currently investigating other potential applications where in vitro monitoring of cell populations is relevant. Over the next few years, the vision is to demonstrate that smart cell scaffolds fabricated with pyrolytic carbon indeed have the potential to become the new black for cell culturing, or at least a complementary method for fundamental studies performed in traditional microwell plates.