Abstract
The misuse of antibiotics in health care has led to increasing levels of drug resistant infections (DRI’s) occurring in the general population. Most technologies developed for the detection of DRI’s typically focus on phenotyping or genotyping bacterial resistance rather than on the underlying cause and spread of DRI’s; namely the misuse of antibiotics. An aptameric based assay has been developed for the monitoring of ampicillin in urine samples, for use in determining optimal antibiotic dosage and monitoring patient compliance with treatment. The fluorescently labelled aptamers were shown to perform optimally at pH 7, ideal for buffered clinical urine samples, with limits of detection as low as 20.6 nM, allowing for determination of ampicillin in urine in the clinically relevant range of concentrations (100 nM to 100 µM). As the assay requires incubation for only 1 h with a small sample volume, 50 to 150 µL, the test would fit within current healthcare pathways, simplifying the adoption of the technology.
Highlights
Antibiotics are used extensively for the vital treatment of bacterial infections in both humans and animals
We demonstrate a diagnostic assay based on fluorescently-labelled aptamers that is capable of quantifying the concentration of ampicillin in clinical urine samples with a high degree of sensitivity at clinical levels
The single stranded DNA (ssDNA) aptamer was selected against ampicillin via the SELEX technique and was confirmed to bind to ampicillin with a high degree of sensitivity and specificity (Kd = 13.4 nM)
Summary
Antibiotics are used extensively for the vital treatment of bacterial infections in both humans and animals. Β-Lactam-based antimicrobials are among the most commonly used classes of antibiotics, showing effectiveness against a wide variety of both Gram-positive and Gram-negative bacteria [4,5,6] Overuse of this important class of antibiotics, partly due to poor dosage management and patients not complying with treatment, has led to the emergence of high levels of resistance [7]. It has been suggested that up to 75% of critically ill patients in intensive care may not be receiving appropriate doses of β-lactam class antibiotics [8] and in excess of 95% of pathogenic Staphylococcus aureus worldwide is resistant to β-lactam antibiotics [7] This high prevalence of resistance has dramatically restricted the use of previously common β-lactam antibiotics. Ampicillin, for example, was the most commonly used β-lactam class antibiotic in healthcare but increasingly, pathogenic
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