The toxicological testing programs of the OECD and the EU contain guidelines which apply to toxicological studies of both, known and newly-developed substances. In recent years, it has become increasingly clear that some substances pose a particular danger to children, and the vulnerability of the developing brain has become the focus of heightened attention. As a result, the OECD and EU now recommend that chemicals with known neurotoxic or teratogenic effects are tested for developmental neurotoxicity. The US Environmental Protection Agency (EPA) also recommends that pesticides should be tested for developmental neurotoxicity. The inclusion of tests for developmental neurotoxicity in the EU's REACH (Registration, Evaluation and Authorization of Chemicals) regulation is currently under debate. These tests will require approx. 30,000 chemicals to be tested for their potential toxicity.
Silicon-based “lab-on-a-chip” technology, developed in the 1980s and 1990s, was initially used in the development of micropumps, microvalves and similar applications. A broad range of applications followed soon, including bioanalysis and the optimization of microfabrication, microlithography and surface structuring technology for DNA separation and cell manipulation in electric fields. Today, the main biological and medical applications of this technology are in the fields of cell culture for biochemical analysis and clinical diagnosis (immunoassays, protein separation and analysis, and polymerase chain reactions (PCRs)). “Lab-on-a-chip” technology is also used in experiments which investigate the electrophysiological and metabolic properties of biological cells, so that suitable cells can be selected for substance screening based on their sensitivity. Combining cell culture with analysis systems creates a new tool that is quick, compact and powerful. This allows online recording of temporal variations in physiological data in dependence of the substances being tested.
Scientists from the Steinbeis Transfer Center for Cell Manipulation and Monitoring Systems (CMMS®) and the Department of Biophysics at the University of Rostock have developed an innovative system for in vitro testing of vertebrate nerve-cell networks. This modular glass-chip system (MOGS) consists of a miniaturized, microstructured glass chip and a preamplifier. The glass chip is the central component, and consists of a 1 mm² multi-electrode array (MEA) with 52 platinum micro-electrodes on a glass support.
The chip has electrical contacts on all four edges, and the MEA input lines on the chip are arranged such that they are as far apart from each other as possible. The chip also includes an interdigital electrode structure (IDES) and a temperature sensor. A small glass trough (Dinner = 8 mm; H = 5 mm) contains the cell culture medium with the test substances. The glass chip is suitable for use with microscopes, and can be steam-sterilized and reused multiple times. Gold spring-pins serve as electrical contacts between the glass chip and a removable adapter board. The preamplifier has a flat shielded casing with a round opening for microscopic observation. Also during electrical network activity measurements. The headstage can be easily connected to a commercial interface for digitalizing the measured values and allowing using suitable analysis software.
The neuron-covered MEA electrodes measure cellular electrical activity. While IDES impedance measurement registers cell adhesion and cell spreading as indicators of cell vitality, and also measures cell spreading on the IDES due to cell growth and proliferation. An integrated sensor monitors the temperature close to the cells and detects any temperature changes. As a next step, the project team plans to integrate additional glass-fiber sensors into the chip to measure acidification and oxygen consumption.
The MOGS can be used in basic research (i.e. for developing stem cell differentiation protocols) and in development and analysis (as a highly sensitive analysis system to detect specific toxic, neurotoxic and developmentally neurotoxic substances in the clinical, environmental, food and pharmaceutical sectors). The system is especially suitable for developing methods to replace animal testing. Metabolic and electrophysiological processes can be monitored online during differentiation of stem cells and neurons on the glass chip. By sowing murine primary cells from a donor organism in several chips simultaneously and processing them in parallel, as in a multi-well system, fewer donor organisms are needed for in vitro testing. The ultimate goal, however, should be to use cell culture lines (containing murine stem cells, for instance) to develop drugs inand investigate the mechanisms of action of chemical substances. MOGS is the first step in this important new direction.
Sebastian Bühler | Marco Stubbe |Philipp Julian Köster |
Dr. Werner Baumann | Prof. Dr. Jan Gimsa
Steinbeis Transfer Center for Cell Manipulation and Monitoring Systems (CMMS)