Gold nanoparticles (AuNP)-based aptasensor for enteropathogenic Escherichia coli detection

Abstract

Diarrhea is a major cause of severe gastrointestinal illness in the infant especially in many developing countries. Although this molecular technique has been accepted as standard technique to detect Diarrhea-causing EPEC, the practical aspect of this technique for in-site rapid screening purposes is still facing a major challenge. In this study, we characterized EPEC specific aptamers and applied it as an AuNP-based aptasensor for point of care (POC) diagnosis purpose. As many as six selected DNA aptamers was screened using target bacteria and the bound aptamer was measured by qPCR technique. Moreover, Kd value for each optimal bound aptamer was measured by using the same technique. Colorimetry assay was applied to test specificity and LOD of AuNP-based aptasensor. Two DNA aptamers have been successfully obtained to detect Enteropathogenic Escherichia coli K.1.1. DNA aptamer S8-7 exhibited constant dissociation (Kd) value of 17.08nM, while DNA aptamer S10-5 exhibited Kd value of 34.14nM. AuNP-based aptasensor showed high selectivity and specificity for EPEC K.1.1 with a limit of detection (LOD) value of 105CFU/mL. Truncation study on DNA aptamer S8-7 showed that elimination of primer binding sequence only slightly increased both performance of detection and LOD value of AuNP-based aptasensor. Further study is necessary to improve AuNP-aptasensor performance such as through mutagenesis approach on targeted DNA aptamers before AuNP-based aptasensor can be applied as a biosensor in point of care (POC) diagnosis.