Aptamer based vanillin sensor using an ion-sensitive field-effect transistor

An aptamer for vanillin was obtained and then used for the development of an aptasensor based on an ion-sensitive field-effect transistor (ISFET). This aptamer (a single-stranded DNA;ssDNA) was selected using the Capture-SELEX protocol, which suites well for selection of aptamers to small molecules. Among six aptamer candidates, the aptamer Van_74 with the highest affinity for vanillin was chosen (elution of 35% of the aptamer from a solid support in the presence of 2 mM of vanillin). Van_74 was characterized using nondenaturating PAGE of washouts from magnetic beads. It is shown that Van_74 binds to vanillin with an dissociation constant of > 7.8 mu M (determined by nondenaturating PAGE) and it was specific to vanillin in comparison with interferents: benzaldehyde, guaiacol, furaneol, ethyl guaiacol and ethyl vanillin. Also it was shown that change of buffer composition greatly affected the binding ability of Van_74. For biosensor fabrication aptamer was immobilised on the Ta2O5-sensitive surface of the ISFET via click-chemistry. Detection scheme implied dehybridisation of the ssDNA probe from the aptamer and release in the solution during the addition of vanillin. As a result, the surface potential increase upon vanillin binding with the aptamer was detected by the transistor. The biosensor had a detection limit of 1.55 x 10(-7) M and a dynamic range from 1.55 x 10(-7) M to 1x10(-6) M. Effective constant K-d,K-eff for vanillin binding on biosensor surface was calculated to be (9 +/- 3) x 10(-7) M. This allows selective detection of vanillin in the mixture of interferents and in samples of coffee extract.