Mian Jiang

Indicator-free electrochemical DNA hybridization biosensor

A new electrochemical hybridization biosensor protocol without an external indicator is described. The biosensor format involves the immobilization of inosine-substituted (guanine-free) probe onto the carbon paste transducer, and a direct chronopotentiometric detection of the duplex formation by the appearance of the guanine oxidation peak of the target. Such a use of the intrinsic DNA electrochemical response for monitoring hybridization events offers several advantages (over the common use of external indicators), including the appearance of a new peak, a flat background, or simplicity. A 4 min short hybridization period allows a detection limit around 120 ng/ml. Performance characteristics of the sensor are described along with future prospects.

New label-free DNA recognition based on doping nucleic-acid probes within conducting polymer films

A new biosensing strategy for in situ electrochemical detection of DNA hybridization is described. The label-free approach relies on the doping of nucleic acid probes within electropolymerized polypyrrole (PPy) films and monitoring the current changes provoked by the hybridization event. The study demonstrates that oligonucleotide probes can serve as the sole counter anion during the growth of conducting PPy films, and maintain their hybridization activity within the host polymer network. Distinct hybridization peak signals, with opposite direction in the presence of complementary and non-complementary DNA sequences, are observed in real time. Such unique response is attributed to concomitant conductivity changes, and offers great promise for DNA diagnostics.

Electrochemical quartz crystal microbalance investigation of surface fouling due to phenol oxidation

Abstract An electrochemical quartz crystal microbalance is employed for investigating the formation of inhibitory polyphenol layers on the surface of gold electrodes. The frequency–time profiles are indicative of the formation of a thin insulating polymeric layer, that meets the rigid film requirement, and are in good agreement with the voltammetric response. Once access to the monomer is blocked, no further frequency change is observed. The passivation process is explored as a function of the specific phenolic compound, its concentration, and operational conditions. Comparison to the charge passed during the film deposition indicates that the efficiency of the polymerization process is dependent upon the phenol concentration. Valuable insights are also gained for the entrapment of glucose oxidase within the electrochemically grown polyphenol.

Real-time monitoring of enzymatic cleavage of nucleic acids using a quartz crystal microbalance

The use of quartz crystal microbalance (QCM) for monitoring in situ the enzymatic cleavage of surface-confined nucleic acids by nucleases is described. Such real-time monitoring of mass changes associated with the enzymatic digestion indicates that the activity and specificity of nucleases is preserved at the gold surface, and can be used for manipulating surface-confined DNAs and RNAs. These observations indicate great promise for using QCM for elucidating the interactions of nucleic acids with enzymes, and for enhancing the power of hybridization biosensors.

Recognition and detection of oligonucleotides in the presence of chromosomal DNA based on entrapment within conducting-polymer networks

Abstract Overlapping voltammetric signals, accrued from redox processes of nucleobases, do not permit discrimination between short oligonucleotides and chromosomal DNA molecules when conventional electrochemical techniques are used. This article describes a new genoelectronic route for discriminating between short oligonucleotides and chromosomal DNA, based on the polypyrrole (PPy) doping process. Such a route relies on the profound effect of short nucleic acid dopants upon the redox activity of PPy, and hence on the square-wave voltammetric signal of the polymer-modified electrode in a blank electrolyte solution. The electropolymeric growth of PPy thus serves for preferential accumulation (by doping) of short oligonucleotides. High selectivity is demonstrated for voltammetric measurements of oligo(dG) 20 and oligo(dT) 20 in the presence of otherwise interfering ss- and ds-DNA. The signals for the oligonucleotides are also not affected by a large excess of chloride or phosphate ions. The response of the new preconcentration (doping)–medium-exchange–voltammetric protocol is proportional to the concentration of the oligonucleotide dopant. Such a new recognition process, based on the doping of conducting-polymer networks, enhances the scope of electroanalysis of nucleic acids.

On-demand electrochemical release of DNA from gold surfaces.

Electrochemical quartz crystal microbalance (EQCM) is used to probe the electrochemically triggered release of nucleic acids from gold surfaces to solutions of physiological pH. The immobilization of nonthiolated DNA onto the gold surface is followed by an electrostatic desorption at -1.0 V (vs. Ag/AgCl). Steady-state frequency signals, corresponding to the removal of 261- and 644-ng/cm2 single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA), respectively, were attained within 75 and 330 s. As expected for electrostatic repulsion, the amount released can be manipulated by tuning the potential. The small nonsteady-state frequency signals observed at lower potentials indicate promise for a sustained DNA release. Applicability to gold ultramicroelectrodes (12.5-microm radius) is demonstrated in connection with voltammetric blocking experiments. We expect that such on-demand electrochemical release would be a useful addition to the arsenal of nonviral gene delivery routes.