Improvement of a highly sensitive and specific whole-cell biosensor by adding a positive feedback amplifier.

ABSTRACT

In this study, we designed a Cd2+ whole-cell biosensor with both positive and negative feedback cascade amplifiers in Pseudomonas putida KT2440 (LTCM) based on our previous design with only a negative feedback amplifier (TCM). The results showed that the newly developed biosensor LTCM was greatly improved compared to TCM. Firstly, the linear response range of LTCM was expanded while the maximum linear response range was raised from 0.05 to 0.1 µM. Meanwhile, adding a positive feedback amplifier further increased the fluorescence output signal of LTCM 1.11-2.64 times under the same culture conditions. Moreover, the response time of LTCM for detection of practical samples was reduced from 6 to 4 h. At the same time, LTCM still retained very high sensitivity and specificity, while its lowest detection limit was 0.1 nM Cd2+ and the specificity was 23.29 (compared to 0.1 nM and 17.55 in TCM, respectively). In summary, the positive and negative feedback cascade amplifiers effectively improved the performance of the biosensor LTCM, resulting in a greater linear response range, higher output signal intensity, and shorter response time than TCM while retaining comparable sensitivity and specificity, indicating better potential for practical applications.