Enhancement of sensitivity in interferometric biosensing by using a new biolinker and prebinding antibody

Recombinant E. coli ACV 1003 (recA::lacZ) was used to measure low concentrations of DNA-darnaging chemicals, which produce β-galactosidase via an SOS regulon system. Very low β-galactosidase activities of less than 0.01 unit/ml, β-galactosidase produced through an SOS response corresponding to the 10 ng/ml (ppb) of DNA damaging chemicals in the environment, can be rapidly determined by using an alternative interferometric biosensor with optically flat thin films of porous silicon rather than by the conventional time-consuming Miller's enzyme assay as well as the ELISA method. In order to enhance the sensitivity in the interferometry, it needs to obtain more uniform distribution and higher biolinking efficiency, whereas interferometric sensing is rapid, cheap, and advantageous in high throughput by using a multiple-well-type chip. In this study, pore size adjusted to 60 rim for the target enzyme β-galactosidase to be bound on both walls of a Si pore and a calyx crown derivative was apllied as a more efficient biolinker. Furthermore, anti-β-galactosidase was previously functionalized with the biolinker for the target β-galactosidase to be specifically bound. When anti-β-galactosidase was bound to the calyx-crown derivative-linked surface, the effective optical thickness was found to be three times as high as that obtained without using anti-β-galactosidase. The resolution obtained was very similar to that afforded by the time-consuming ELISA method; however, the reproducibility was still unsatisfactory, below 1 unit β-galactosidase/ ml, owing to the microscopic non-uniform distribution of the pores in the etched silicon surface.