Real-Time Intracellular Analysis of Kanamycin Using Microaptasensors

With the emergence of multidrug-resistant bacteria, infection-related death toll is on the rise. Overuse of antibiotics and their leakage into waterways have transformed the environment into a sink, resulting in bacterial resistance permeating through all tiers of the food cycle. As one of the primary sources of food, fish and fish products such as fish eggs must be studied for their ability to accumulate relevant antibiotics. While the accumulation of these pharmaceuticals has previously been studied, there remains a need to analyze these processes in real time. Electrochemical aptamer-based sensor technology allows for selective, real-time monitoring of small molecules. Herein, we report the first use of miniaturized electrochemical aptamer-based sensors for the analysis of the passive uptake of the aminoglycoside antibiotic, kanamycin, in single salmon eggs. We use pulled platinum microelectrodes and increase the surface area at the electrode tip through dendritic gold deposition. These electrodes showed a 100-fold increase in DNA immobilization on the electrode surface as compared to bare microelectrodes. Additionally, the sensors showed improved stability in complex biological media over an extended period of time when compared to the more widely used macrosensors (r = 1 mm). The sensor range was determined to extend from nanomolar to micromolar concentrations of kanamycin in fish egg lysate and when used in a single salmon egg the μ-aptasensors were able to monitor the passive uptake of kanamycin over time. The accumulation kinetics were simulated using COMSOL Multiphysics software. This research presents the first reported record of passive uptake of a small molecule in a single cell in real-time using electrochemistry.