title: Analyte-driven switching of DNA charge transport: de novo creation of electronic sensors for an early lung cancer biomarker.
abstract: A general approach is described for the de novo design and construction of aptamer-based electrochemical biosensors, for potentially any analyte of interest (ranging from small ligands to biological macromolecules). As a demonstration of the approach, we report the rapid development of a made-to-order electronic sensor for a newly reported early biomarker for lung cancer (CTAP III/NAP2). The steps include the in vitro selection and characterization of DNA aptamer sequences, design and biochemical testing of wholly DNA sensor constructs, and translation to a functional electrode-bound sensor format. The working principle of this distinct class of electronic biosensors is the enhancement of DNA-mediated charge transport in response to analyte binding. We first verify such analyte-responsive charge transport switching in solution, using biochemical methods; successful sensor variants were then immobilized on gold electrodes. We show that using these sensor-modified electrodes, CTAP III/NAP2 can be detected with both high specificity and sensitivity (K(d) ~1 nM) through a direct electrochemical reading. To investigate the underlying basis of analyte binding-induced conductivity switching, we carried out Förster Resonance Energy Transfer (FRET) experiments. The FRET data establish that analyte binding-induced conductivity switching in these sensors results from very subtle structural/conformational changes, rather than large scale, global folding events. The implications of this finding are discussed with respect to possible charge transport switching mechanisms in electrode-bound sensors. Overall, the approach we describe here represents a unique design principle for aptamer-based electrochemical sensors; its application should enable rapid, on-demand access to a class of portable biosensors that offer robust, inexpensive, and operationally simplified alternatives to conventional antibody-based immunoassays.
authors: Department of Chemistry, Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada.
authors: Jason M
authors: JM
authors: Thomas
authors: Banani
authors: B
authors: Chakraborty
authors: Dipankar
authors: D
authors: Sen
authors: Hua-Zhong
authors: HZ
authors: Yu
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry*; D001483:Base Sequence; D014408:Biomarkers, Tumor / Q000032:analysis*; D015374:Biosensing Techniques / Q000191:economics / Q000379:methods*; D004247:DNA / Q000737:chemistry; D055664:Electrochemical Techniques / Q000191:economics / Q000379:methods; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008175:Lung Neoplasms / Q000175:diagnosis*; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis*; D052156:SELEX Aptamer Technique; D012680:Sensitivity and Specificity
chemical_list: D052157:Aptamers, Nucleotide; D014408:Biomarkers, Tumor; D010455:Peptides; C009862:connective tissue-activating peptide; D004247:DNA