Abstract
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.
Highlights
DNA amplification is an essential step in most nucleic acid-based diagnosis methods, including those used in the diagnosis of infectious diseases
In real-time Polymerase chain reaction (PCR), which is known as quantitative PCR
Disadvantages of real-time PCR include the need for expensive reagents and optical detection apparatus, as well as the need for specialist knowledge for designing specific DNA probes
Summary
DNA amplification is an essential step in most nucleic acid-based diagnosis methods, including those used in the diagnosis of infectious diseases. Polymerase chain reaction (PCR), involving the amplification of specific DNA sequences from single molecules using multiple enzymatic reaction cycles, is the best-known DNA amplification method, others, including isothermal amplification methods, have been developed [1]. In real-time PCR, which is known as quantitative PCR (qPCR), optical methods are used to measure amplicon levels after each cycle. Real-time PCR has a broad dynamic range of 7–8 logarithmic decades, and is useful for quantitative measurements [2,3,4]. Disadvantages of real-time PCR include the need for expensive reagents and optical detection apparatus, as well as the need for specialist knowledge for designing specific DNA probes.
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