5 Conclusion
The cell-free system with unique advantages provides a novel design platform for the development of biosensors. Considering the openness and rapid prototyping capability of CFPS, the CFPS system can open up more design space for the construction and optimization of biosensors and genetic circuits. Cell-free biosensors based on TF, RNA, and toehold switch can reduce the number of cycles required to design and help build complex genetic networks. CFPS system can also be integrated with mathematical modeling and high technologies (e.g., automation and microfluidics) to unlock the potential of cell-free biosensors.
In the practical applications of cell-free biosensors, the lyophilization technology based on the portable paper platform can maintain the stability of the system, which promotes its portable use and reduces the cost. However, there are still many challenges to be addressed. The decline of the activity after the storage is a problem that needs to be further improved. To improve in situ detection, multiplexing functions, the sensitivity, and the specificity of cell-free biosensors need to be further optimized.
In addition, cell-free biosensors can use CFPS systems other than E. coli , such as yeast and mammalian cells, to develop more sophisticated biosensors. By improving the sensor recognition mechanisms and combining cell-free systems with other materials (e.g., silicon), more types of functional cell-free biosensors can be developed. It might expand the detection range of cell-free biosensors to sense the odor, air, temperature, light, and osmotic pressure. With further evolution of cell-free biosensors, besides environmental monitoring and human health diagnosis, they will increasingly expand their applications to food testing, classroom education, and others, making them more practical and commercial.