Glucose-loaded liposomes for amplified colorimetric immunoassay of streptomycin based on enzyme-induced iron(II) chelation reaction with phenanthroline

This work designed a signal-on competitive-type colorimetric immunoassay of streptomycin (STR) using glucose-loaded liposome (GLL) and unique iron(II)-phenanthroline [Fe(II)-Phen] system. The assay was executed based on the change in the color and absorbance of Fe(II)-Phen system, followed by enzyme-triggered Fe(II) oxidization into Fe(III) in the presence of glucose oxidase (GOx). To construct such an assay, STR-bovine serum albumin (STR-BSA) conjugates were covalently linked with glucose-loaded liposome, whereas anti-STR antibodies were coated onto a microplate. In the presence of STR, a competitive immunoreaction was executed between the analyte and STR-BSA-GLL for the immobilized anti-STR. Upon addition of Triton X-100, the carried liposome was dissociated to release the loaded glucose, which was oxidized by GOx into gluconic acid and hydrogen peroxide (H2O2). The as-generated H2O2 oxidized iron(II) into iron(III), and decreased the complexation of iron(II) with Phen, thereby resulting in the color change and the decreasing absorbance. Under optimal conditions, the colorimetric immunoassay with the Fe(II)-Phen system exhibited a low detection limit (LOD) of 0.4 pg mL−1 with a wide dynamic working range from 1.0 pg mL−1 to 20 ng mL−1 STR, accompanying good reproducibility and high specificity.