DNA Immobilization and Hybridization Detection by the Intrinsic Molecular Charge Using Capacitive Field-Effect Sensors Modified with a Charged Weak Polyelectrolyte Layer

Abstract

Miniaturized setup, compatibility with advanced micro- and nanotechnologies, and ability to detect biomolecules by their intrinsic molecular charge favor the semiconductor field-effect platform as one of the most attractive approaches for the development of label-free DNA chips. In this work, a capacitive field-effect EIS (electrolyte-insulator-semiconductor) sensor covered with a layer-by-layer prepared, positively charged weak polyelectrolyte layer of PAH (poly(allylamine hydrochloride)) was used for the label-free electrical detection of DNA (deoxyribonucleic acid) immobilization and hybridization. The negatively charged probe single-stranded DNA (ssDNA) molecules were electrostatically adsorbed onto the positively charged PAH layer, resulting in a preferentially flat orientation of the ssDNA molecules within the Debye length, thus yielding a reduced charge-screening effect and a higher sensor signal. Each sensor-surface modification step (PAH adsorption, probe ssDNA immobilization, hybridization with complementary target DNA (cDNA), reducing an unspecific adsorption by a blocking agent, incubation with noncomplementary DNA (ncDNA) solution) was monitored by means of capacitance-voltage and constant-capacitance measurements. In addition, the surface morphology of the PAH layer was studied by atomic force microscopy and contact-angle measurements. High hybridization signals of 34 and 43 mV were recorded in low-ionic strength solutions of 10 and 1 mM, respectively. In contrast, a small signal of 4 mV was recorded in the case of unspecific adsorption of fully mismatched ncDNA. The density of probe ssDNA and dsDNA molecules as well as the hybridization efficiency was estimated using the experimentally measured DNA immobilization and hybridization signals and a simplified double-layer capacitor model. The results of field-effect experiments were supported by fluorescence measurements, verifying the DNA-immobilization and hybridization event.

Capacitive Field-Effect EIS Chemical Sensors and Biosensors: A Status Report

“…In recent years, DNA biosensors have been increasingly recognized as powerful tools in many fields of application, including molecular diagnostics, pathogen identification, drug screening, food safety, forensic and parental testing, or detecting biowarfare agents. The vast majority of DNA-modified EIS (DNA-EIS) biosensors reported in the literature is based on detecting a DNA-hybridization reaction [ 7 , 27 , 135 , 136 , 137 , 138 ], although the detection of single-stranded DNA (ssDNA) [ 8 , 132 ] and double-stranded DNA (dsDNA) [ 132 , 139 , 140 , 141 ], as well as other DNA-recognition events, like single-base mismatch [ 130 ], the by-product (protons) of the nucleotide base incorporation reaction [ 36 ], and DNA amplification by polymerase chain reaction (PCR) [ 139 , 140 , 142 , 143 , 144 ], have been demonstrated as well.…”

Hybrid Layer‐by‐Layer Film of Polyelectrolytes‐Embedded Catalytic CoFe2O4 Nanocrystals as Sensing Units in Capacitive Electrolyte‐Insulator‐Semiconductor Devices

“…The potential of this method rises from the variety of types of materials (organic and inorganic) that can be used, as well as, the simple experimental procedure to assemble nanostructured films with synergy among the materials . In particular, the LbL method has been demonstrated to be advantageous for incorporating nanofilms onto EIS sensors, proving to be suitable to study proof‐of‐concept sensing units for biomedical applications . Previous reports have proved that the use of LbL films is beneficial to preserve the enzyme activity, as well as, to enhance the output signal and sensitivity of enzymatic EIS sensors .…”

“…Previous reports have proved that the use of LbL films is beneficial to preserve the enzyme activity, as well as, to enhance the output signal and sensitivity of enzymatic EIS sensors . Furthermore, the use of polyelectrolytes in LbL films as sensing membrane have demonstrated to be appropriate for the detection of charged macromolecules …”

“…Particularly for the latter, it is reported that compound‐based MFe 2 O 4 (transition metal) presents propitious properties for H 2 O 2 determination due to its spinel structures, as M x+ ions may interact with the O–O bond of H 2 O 2 , releasing ions and electrons that can be measured by using different electrochemical methods . In this sense, the capacitive EIS sensor can be an alternative sensing platform toward H 2 O 2 detection, as the release of H + ‐ions in solution can promote an interface potential across the solid–liquid junction (sensor chip–H 2 O 2 solution), which can be detected by the EIS sensor . However, to our knowledge there are no studies reported involving EIS sensors using CoFe 2 O 4 as sensitive layer for H 2 O 2 detection.…”

Development and Bio-imaging Applications of a Chemical Imaging Sensor

“…In the measurement of charged DNA molecules using an LAPS or EIS capacitive sensor, (81)(82)(83) a low-ionic-strength environment was used to mitigate the shielding effect. For further applications of a chemical imaging sensor to cell measurement, it is necessary to overcome the problem of the shielding effect.…”