Abstract
For the analysis of massive samples containing multiple analytes, the enhancement of detection efficiency is crucial. In this study, a facile method was developed for sequential detection of chloramphenicol (CAP) and gentamicin (GEN) in complex samples, e.g. milk, using a surface plasmon resonance (SPR)-based biosensor. Based on the immune inhibition format, two conjugates of antigen and bovine serum albumin (BSA)—denoted as CAP–BSA and GEN–BSA—were grafted on the same channel of the SPR sensor chip. Two standard curves for CAP and GEN were separately obtained by first mixing a single antibody with different concentrations of the relevant antigen. Moreover, different regeneration solutions were screened for sequential analysis. An alkaline solution was found to completely remove the antibody against GEN (AbGEN) from the chip, but it exhibited limited ability to dissociate the antibody against CAP (AbCAP). Therefore, alkaline solution and Gly-HCl solutions are successively applied to elute AbGEN and AbCAP, respectively. By gradual elutions, CAP and GEN concentrations were simultaneously calculated with limit of detection values of 5.28ng/mL and 2.26ng/mL, respectively. Furthermore, the spiking milk samples with CAP and GEN validated the assay with recoveries of 77.6–101.1%. Therefore, this method is expected to improve the detection efficiency of SPR biosensors.
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