In the development of modern biotechnology, microbioreactors play a key role in high-throughput screening of bacterial strains and small-scale culture experiments. With the continuous innovation in the development of bioreactor process technology, various micro bioreactors have brought more development space to biopharmaceuticals, industrial production and other fields.
Microbioreactor (MBR) refers to small-scale bioreactors with a volume of less than a few hundred milliliters and miniature bioreactors with a volume of less than 100 mL. Micro bioreactors can often carry out several or even dozens of parallel culture processes at the same time, so they have certain high-throughput characteristics.
At present, MBR has been effectively used in the identification of strain characteristics in the strain screening process, and the rapid optimization of microbial and animal cell culture media and culture processes, and has become one of the important development trends of bioreactors.
The concept of MBR can be mainly divided into two categories:
First, from top to bottom, traditional bioreactors are reduced, integrated and arrayed to provide more process information and throughput;
The second is to work from the bottom up and configure process detection devices (such as dissolved oxygen and pH detection) on traditional high-throughput devices (such as shake flasks and microplates) to provide certain process information.
Typical MBRs include microflow reactors, orifice reactors, shake flask reactors, stirred reactors, etc., and their working volumes are microliters, milliliters, tens of milliliters, tens to hundreds of milliliters, respectively.
At present, the main purpose of miniaturization and miniaturization of bioreactors is to conduct high-throughput experiments to speed up the research and development process and shorten the time to market. Achieving this goal relies on suitable materials and processing technologies, compact and reliable sensors, actuators that control process parameters in tiny volumes, and supporting automation and data processing software.
•Materials and processing
Compatibility of materials and reaction systems is the first issue to consider. Even if it is chemically compatible, many biological reaction products or substrates are adsorbed to the surface of the material, which will affect the culture process and result analysis. This problem is particularly prominent due to the relatively large specific surface area of micro-reactors.
In addition, low-quality materials may also contain heavy metals and other substances that are harmful to cells or microorganisms, affecting growth and product production. Some trace metals may also act as catalysts. Since the process parameters of micro-scale reactors, such as biomass concentration, pH, dissolved oxygen, etc., are mostly measured optically through the container wall, the light transmission performance of the material cannot be ignored.
In order to enable rapid mass production, the materials used in micro-scale reactors need to be easy to process. All things considered, high-purity polystyrene is the preferred material for micro-scale bioreactors, and the most economical production method is injection molding. For more complex microstructures, they need to be further processed or assembled through other processes after injection molding.
•Sensor Technology
The measurement and control of process parameters such as dissolved oxygen, pH, carbon dioxide, glucose, and temperature are the most critical aspects of micro-bioreactor technology and were once the bottleneck restricting its development.
Non-invasive optical sensors are the best choice for micro-bioreactors, which can continuously measure and record the changing trends of parameters throughout the fermentation process without sampling.
❶Dissolved oxygen
The more mature dissolved oxygen measurement applied in micro bioreactors is based on the quenching effect of oxygen on fluorescence, that is, the phenomenon that the luminous intensity of fluorescent substances decays and accelerates under the action of oxygen atoms. In the 1980s, people discovered that the stability and sensitivity of fluorescent substance particles fixed in a silica gel film were greatly improved under the protection of the film. Since then, optical dissolved oxygen sensors based on this principle have been widely used, and are not limited to micro-sized ones. Bioreactor.
❷PH
Typical optical pH sensors rely on the property of specific pigments to absorb or emit fluorescence when protonated or deprotonated. The fluorescence intensity emitted by these pigments generally decreases with increasing pH. The area of good linear range is approximately 1 pH unit on either side of the pKa. Beyond this range, the signal-to-noise ratio will be severely affected. Therefore, when selecting a disposable micro-bioreactor, the adaptation of the pH range is particularly important.
❸Detection of other parameters
In addition to direct pH measurements, pH-sensitive pigments are also used to indirectly measure dissolved CO2 based on bicarbonate dissociation equilibrium.
The most mature technology for glucose measurement is based on enzymatic catalysis, including standard-sized electrodes and micro-miniature products integrated into culture vessels. This electrode itself will consume glucose and will have a certain impact on the microreactor.
Temperature measurement and control of micro-scale reactors is important in two aspects. In addition to the requirements of the biochemical reaction itself, temperature also determines the accuracy of other sensing elements.
Manual liquid filling, inoculation, sampling and other operations on micro bioreactors are almost impossible to complete effectively. Therefore, these equipment are generally equipped with additional liquid reagent processing systems, such as integrated liquid reagent processing systems and external automatic sampling systems.
These devices are essentially industrial robots. With the help of experimental design software, experimenters do not need to perform manual calculations. They only need to simply enter the range and specify the number of reactors to be used, and the system can automatically complete the configuration and timing of parameters such as initial substrate concentration. sampling.
At the same time, the control software supporting micro bioreactors is increasingly integrating data visualization and analysis functions. This is compared with the human-computer interaction interface developed based on programmable controllers or embedded systems for traditional desktop bioreactors. Much more convenient and faster.