In 2005, our lab invented the micrometer-sized reactor array, termed “femtoliter chamber array” . It was aimed to quantify the extremely small amount of ATP produced from a single F1 motor protein by forcibly reversing it with magnetic tweezer systems. The advantage of femtoliter chamber is in the condensation of ATP to enable its detection. Confining ATP molecules in an extremely small reaction volume, comparable to that of a single E. coli, concentrates ATP to detectable levels .
The femtoliter chamber array was also applied to single-molecule enzymatic assay. Individual enzyme molecules are encapsulated by a fluorogenic substrate in the reactors. After few minutes of incubation, the chambers containing the single enzyme molecule give bright fluorescent signal whereas the empty ones do not. By counting the number of bright spots, we can quantify the amount of enzyme. We termed this method ‘Digital Bioassay’. This method is quite simple, yet versatile. One of the representative digital bioassays that is expected to be used widely for diagnostic purpose is ‘Digital ELISA’ .
We are also in the process of expanding the applications of Digital Bioassays such as in multicolor digital counting , digital viral counting, and digital isothermal amplification. 1. Y. Rondelez, et al., 2005 Nature 433, 773-777
2. Y. Rondelez, et al., 2005 Nat. Biotech. 3, 361-365
3. S.H. Kim, et al., 2012 Lab Chip 12, 4986-4991
4. Y. Obayashi, et al., 2015 Analyst 140, 5065-5073