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: Jason M Thomas, Banani Chakraborty, Dipankar Sen, Hua-Zhong Yu
• Journal: Journal of the American Chemical Society (J Am Chem Soc) Vol. 134 Issue 33 Pg. 13823-33 (Aug 22 2012) ISSN: 1520-5126 [Electronic] United States
• PMID: 22835075 (Publication Type: Evaluation Study, Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Aptamers, Nucleotide
• Biomarkers, Tumor
• Peptides
• connective tissue-activating peptide
• DNA

Topics

• Aptamers, Nucleotide (chemistry)
• Base Sequence
• Biomarkers, Tumor (analysis)
• Biosensing Techniques (economics, methods)
• DNA (chemistry)
• Electrochemical Techniques (economics, methods)
• Fluorescence Resonance Energy Transfer
• Humans
• Lung Neoplasms (diagnosis)
• Molecular Sequence Data
• Peptides (analysis)
• SELEX Aptamer Technique
• Sensitivity and Specificity