Detection of PCR-Amplified Tuberculosis DNA Fragments with Polyelectrolyte-Modified Field-Effect Sensors

Abstract

Field-effect-based electrolyte-insulator-semiconductor (EIS) sensors were modified with a bilayer of positively charged weak polyelectrolyte (poly(allylamine hydrochloride) (PAH)) and probe single-stranded deoxyribonucleic acid (ssDNA) and are used for the detection of target complementary deoxyribonucleic acid (cDNA) in different test solutions. The sensing mechanism is based on the detection of the intrinsic molecular charge of target cDNA molecules after the hybridization event between cDNA and immobilized probe ssDNA. The test solutions contain synthetic cDNA oligonucleotides (with a sequence of tuberculosis mycobacteria genome) or PCR-amplified DNA (which origins from a template DNA strand that has been extracted from M. avium paratuberculosis-spiked human sputum samples), respectively. Sensor responses up to 41 mV have been measured for the test solutions with DNA, while only small signals of ~5 mV was detected for solutions without DNA. The lower detection limit of the EIS sensors was ~0.3 nM and the sensitivity was ~7.2 mV/decade. Fluorescence experiments using SybrGreen I fluorescence dye support the electrochemical results.