Determination of Shigella flexneri by a Novel Fluorescent Aptasensor

Abstract

A simple, sensitive, selective, and homogeneous fluorescent aptasensor was constructed for Shigella flexneri by using a dye-labeled aptamer and graphene oxide using target recycling amplification. Unique recognition sequences of restriction endonuclease ApaI from E. coli that carried the cloned apaIR gene from Acetobacter pasteurianus were incorporated into a single-stranded signal probe labeled with carboxyfluorescein at the 5′ and 3′ ends. In the absence of target bacteria, probes were adsorbed and quenched by graphene oxide. After the addition of the analyte, an iterative process called target recycling amplification was triggered. Specific binding of the aptamer to the target liberated probes from graphene oxide. The associated probes were recognized and cleaved by the enzyme. In addition, the released targets were reemployed in subsequent measurements. Under the optimum conditions, a linear calibration relationship was displayed from 500 to 109 CFU/mL with a limit of detection of 100 CFU/mL and high selectivity. Consequently, this aptasensor is an attractive alternative to conventional methodologies.

Keywords:

• Fluorescent aptasensor
• Graphene oxide
• Restriction endonuclease ApaI
• Shigella
• Target recycling