Rapid and sensitive detection of Nampt (PBEF/visfatin) in human serum using an ssDNA aptamer-based capacitive biosensor

Official URL: http://dx.doi.org/10.1016/j.bios.2012.05.036

A single-stranded DNA (ssDNA) aptamer was successfully developed to specifically bind to nicotinamide phosphoribosyl transferase (Nampt) through systematic evolution of ligands by exponential enrichment (SELEX) and successfully implemented in a gold-interdigitated (GID) capacitor-based biosensor. Surface plasmon resonance (SPR) analysis of the aptamer revealed high specificity and affinity (Kd=72.52 nM). Changes in surface capacitance/charge distribution or dielectric properties in the response of the GID capacitor surface covalently coupled to the aptamers in response to changes in applied AC frequency were measured as a sensing signal based on a specific interaction between the aptamers and Nampt. The limit of detection for Nampt was 1 ng/ml with a dynamic serum detection range of up to 50 ng/ml; this range includes the clinical requirement for both normal Nampt level, which is 15.8 ng/ml, and Nampt level in type 2 diabetes mellitus (T2DM) patients, which is 31.9 ng/ml. Additionally, the binding kinetics of aptamer–Nampt interactions on the capacitor surface showed that strong binding occurred with increasing frequency (range, 700 MHz–1 GHz) and that the dissociation constant of the aptamer under the applied frequency was improved 120–240 times (Kd=0.3–0.6 nM) independent on frequency. This assay system is an alternative approach for clinical detection of Nampt with improved specificity and affinity.