A label-free biosensor for selective detection of Gram-negative bacteria based on the oxidase-like activity of cupric...

A label-free biosensor based on cupric oxide (CuO) nanoparticles was constructed for the selective detection of Gram-negative bacteria. CuO possesses oxidase-like activity and can catalyze the oxidation of o-phenylenediamine (OPD) to produce oxidized OPD, which has a fluorescence emission at 573 nm under excitation at 423 nm. The mechanism study suggests that the oxygen vacancies of CuO can activate the dissolved oxygen to form superoxide anions, which in turn oxidize OPD. Gram-negative bacteria can reduce part of Cu(II) in CuO to Cu(I) based on their copper homeostasis system, thus inhibiting the oxidation of OPD and decreasing the fluorescence intensity of the catalytic system. This principle was utilized to construct a biosensor to realize the selective detection of Gram-negative bacteria successfully. The biosensor exhibited a good linear correlation toward the logarithm concentration of three Gram-negative bacteria with R² ≥ 0.985. It was applied to detect three Gram-negative bacteria in eggshell, Chinese cabbage, and the Pearl River water samples, with recoveries ranging from 92.4 to 107%. Moreover, a smartphone-based portable device was designed and fabricated to realize the on-site detection of bacteria. The results of the portable device were comparable to those of fluorescence spectrophotometry, suggesting that the portable device has tremendous potential in the on-site detection of bacteria.