Zhenhao Ding

Rapid microbead-based DNA detection using dielectrophoresis and impedance measurement

Polymerase chain reaction (PCR) is a powerful tool for diagnostic procedures in bacterial and viral infections. The authors propose a new electrical technique for rapid detection of DNA amplified by PCR using dielectrophoresis (DEP) of microbeads. The method is based on dramatic alteration of DEP characteristics of microbeads caused by DNA labeling. DNA-labeled microbeads are trapped on a microelectrode under the action of positive DEP, whereas pristine ones are not. DEP-trapped microbeads are measured impedimetrically to realize rapid and quantitative detection of the amplified DNA. The validity of the proposed method was demonstrated by detection of PCR-amplified DNA of viruses.

Dielectrophoresis and dielectrophoretic impedance detection of adenovirus and rotavirus

The aim of this study is the electrical detection of pathogenic viruses, namely, adenovirus and rotavirus, using dielectrophoretic impedance measurement (DEPIM). DEPIM consists of two simultaneous processes: dielectrophoretic trapping of the target and measurement of the impedance change and increase in conductance with the number of trapped targets. This is the first study of applying DEPIM, which was originally developed to detect bacteria suspended in aqueous solutions, to virus detection. The dielectric properties of the viruses were also investigated in terms of their dielectrophoretic behavior. Although their estimated dielectric properties were different from those of bacteria, the trapped viruses increased the conductance of the microelectrode in a manner similar to that in bacteria detection. We demonstrated the electrical detection of viruses within 60 s at concentrations as low as 70 ng/ml for adenovirus and 50 ng/ml for rotavirus.

DNA detection using microbeads-based dielectrophoretic impedance measurement

A new electrical technique for rapid detection of DNA amplified by polymerase chain reaction (PCR) using dielectrophoresis (DEP) of microbeads was demonstrated. The amplicons were chemically immobilized on the microbeads. The DNA immobilization on the microbeads alters their DEP behavior in such a way that they are trapped on a microelectrode under the action of positive DEP, whereas pristine microbeads are not. Combining the dramatic alteration in DEP characteristics with impedance measurement leads to rapid and quantitative detection of the amplicon in a few seconds. The proposed microbead-based assay could be applicable for rapid, quantitative, and automated detection of virus infection.

Bacterial detection based on polymerase chain reaction and microbead dielectrophoresis characteristics

In this study, an electrical DNA detection method was applied to bacterial detection. DNA was extracted from bacteria and amplified by polymerase chain reaction. The microbeads were labelled with amplicons, altering their surface conductance and therefore their dielectrophoresis characteristics. Amplicon-labelled microbeads could thus be trapped within a high-strength electric field, where they formed a pearl chain between the electrodes, resulting in an increased conductance between the electrodes. This method reduces the amplicon detection time from 1-2 h to 15 min, compared with the conventional method. The presented method realised quantitative detection of specific bacteria at concentrations above 1 × 105 and 2.4 × 104 CFU/ml for bacterial solutions with and without other bacterial presence, respectively.

Effect of DNA length on dielectrophoretic characteristics of DNA-labeled microbeads

Polymerase chain reaction (PCR) is a powerful tool for diagnostic procedures in bacterial and viral infections. We have developed a new electrical technique for rapid detection of DNA amplified by PCR using dielectrophoresis (DEP) of microbeads that are chemically labeled with the amplicons. The DNA immobilization on the microbeads alters their DEP behavior in such a way that they are trapped on a microelectrode under the action of positive DEP, whereas pristine microbeads are not. Combining the dramatic alteration in DEP characteristics with impedance measurement leads to rapid and quantitative detection of amplicons. The method is based on the surface conductivity dependence of microbeads DEP characteristics. It was expected that the surface conductivity would depend on the length of DNA fragments immobilized on a microbeads. In this study, it was found that the crossover frequency was dependent on the length of DNA.

DNA detection microfluidic device based on negative dielectrophoresis of DNA labeled microbeads

In this study, we focus on the alteration of negative dielectrophoresis (n-DEP) caused by small amount of DNA labeling on the microbeads and propose a new design of microfluidic device for more sensitive DNA detection. In previous study, we proposed a new electrical technique for rapid detection of DNA amplified by polymerase chain reaction (PCR). The method is based on dramatic alteration of dielectrophoresis (DEP) characteristics of microbeads caused by DNA labeling. The DNA labeling of microbeads alters their DEP behavior in such a way that they are trapped on a microelectrode under the action of positive DEP (p-DEP), whereas bare microbeads are not, due to the n-DEP. However, this method requires at least 105 copies of DNA labelling on a microbead to alter the DEP behavior of microbeads from negative to positive. As a result, we are able to distinguish bare microbeads and microbeads labeled with DNA at DNA to microbead ratio of 3×103: 1 and 3×104: 1, based on the n-DEP alteration.

Comparison of Sensitivity and Quantitation between Microbead Dielectrophoresis-Based DNA Detection and Real-Time PCR.

In this study, we describe a microbead-based method using dielectrophoresis (DEP) for the fast detection of DNA amplified by polymerase chain reaction (PCR). This electrical method measures the change in impedance caused by DEP-trapped microbeads to which biotinylated target DNA molecules are chemically attached. Using this method, measurements can be obtained within 20 min. Currently, real-time PCR is among the most sensitive methods available for the detection of target DNA, and is often used in the diagnosis of infectious diseases. We therefore compared the quantitation and sensitivity achieved by our method to those achieved with real-time PCR. We found that the microbead DEP-based method exhibited the same detection limit as real-time PCR, although its quantitative detection range was slightly narrower at 10–105 copies/reaction compared with 10–107 copies/reaction for real-time PCR. Whereas real-time PCR requires expensive and complex instruments, as well as expertise in primer design and experimental principles, our novel method is simple to use, inexpensive, and rapid. This method could potentially detect viral and other DNAs efficiently in combination with conventional PCR.

Rapid size determination of PCR amplified DNA by beads-based dielectrophoretic impedance spectroscopy

This paper reports a new method to determine the size of DNA amplified by polymerase chain reaction (PCR). After PCR, the amplified DNA was attached on dielectric microbeads, then, the DNA labeled microbeads were trapped on a microelectrode by dielectrophoresis (DEP). The size of the DNA was determined by measuring frequency dependent impedance of the trapped microbeads as an impedance spectroscopy. The size of various DNAs, 142, 204, 391, 796 bp, were successfully identified from the impedance spectrograms. This method determines DNA size in 20 min after PCR.

Dielectrophoretic characteristics of microbeads labeled with DNA of various lengths

Polymerase chain reaction (PCR) is one of the most sensitive and specific detection methods of bacterial and viral infections. The authors proposed a new electrical technique for rapid detection of DNA amplified by PCR using dielectrophoresis (DEP) of microbeads. The method is based on dramatic alteration of DEP characteristics of microbeads caused by DNA labelling. DNA labeled microbeads are trapped on a microelectrode under the action of positive DEP, whereas pristine microbeads are not. DEP-trapped microbeads can be measured impedimetrically to realize rapid and quantitative detection of the amplified DNA. In this study, it was aimed to reveal how DNA length affects DEP characteristic of DNA-labeled microbeads. Dielectrophoretic crossover from the negative to the positive was measured for microbeads labeled with DNA length in 204 bp, 391 bp and 796 bp. After theoretical fitting of DEP crossover data, it was revealed that the surface conductance increased when the length of labeled DNA increased.

Sensitive and quantitative DNA detection by beads-based dielectrophoretic impedance measurement

We proposed that a rapid detection method of DNA amplified by polymerase chain reaction (PCR) using dielectrophoresis (DEP) of microbeads. This electrical method measures the impedance change of the DEP trapped microbeads on which target DNA molecules are chemically attached. The method realizes easy and rapid DNA detection after conventional PCR. In this study, it was demonstrated that quantitative and high sensitivity detection of virus genome using the method as comparing with real-time PCR. Real-time PCR detects target DNA sensitive and quantitative by real-time monitoring of DNA amplification. As the results, the DNA detection method using microbeads DEP showed the same detection limit as the real-time PCR, and demonstrated the quantitative detection in the range of 101 to 105 copies/reaction. Our new method can realize inexpensive and rapid PCR-based viral detection by combined with conventional PCR.