Characterization and analysis of real-time capillary convective PCR toward commercialization

Qiu, Xianbo; Zhang, Shiyin; Mei, Lanju; Wu, Di; Guo, Qi; Li, Ké; Ge, Shengxiang; Ye, Xiangzhong; Xia, Ningshao; Mauk, Michael G.

doi:10.1063/1.4977841

Cited by 16 publications

(12 citation statements)

References 40 publications

(38 reference statements)

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“…Previously, we successfully developed a couple of CPCR systems for rapid nucleic acid amplification within 30 min, which all relied on a capillary tube reactor (inner and outside diameter: 1.6 mm and 3.3 mm) to introduce consistent thermal convection with pseudo-isothermally heating [21][22][23][24]. The relationship of the sample/reaction volume vs. thermal characteristics of the CPCR reactor was found in our previously published study [26].…”

Section: Single Cpcr Reactor With Sample Preparationmentioning

confidence: 93%

“…For a properly designed CPCR reactor, a consistent temperature field on the reactor must be established to achieve the stable and repeatable thermal convection for robust amplification [19,20]. Compared to conventional PCR, which normally takes 1-2 h, CPCR can be performed within 30 min, which is competitive in POC diagnostics [21][22][23][24]. Recently, few microfluidic CPCR NAAT systems with complicated sample preparation have been reported [25], which nevertheless demonstrate their potentiality in POC diagnostics with simple heating and ultra-fast amplification.…”

Section: Introductionmentioning

confidence: 99%

“…So far, portable microfluidic NAAT systems have not become widespread, and improvements are needed in performance, cost, and ease-of-use. Based on our experience on CPCR [21][22][23][24], by incorporating sample preparation, this work aims to demonstrate an integrated, simple, low-cost, portable, microfluidic CPCR NAAT system with simplified instrumentation. For simplified sample preparation, the CPCR reactor is integrated with a PCR compatible solid phase membrane on the bottom for nucleic acid extraction, which provides the nucleic acid templates without elution.…”

Section: Introductionmentioning

confidence: 99%

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An Integrated, Real-Time Convective PCR System for Isolation, Amplification, and Detection of Nucleic Acids

Miao

Guo

et al. 2022

Chemosensors

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Convective PCR (CPCR) can perform rapid nucleic acid amplification by inducing thermal convection to continuously, cyclically driving reagent between different zones of the reactor for spatially separate melting, annealing, and extending in a capillary tube with constant heating temperatures at different locations. CPCR is promoted by incorporating an FTA membrane filter into the capillary tube, which constructs a single convective PCR reactor for both sample preparation and amplification. To simplify fluid control in sample preparation, lysed sample or wash buffer is driven through the membrane filter through centrifugation. A movable resistance heater is used to heat the capillary tube for amplification, and meanwhile, a smartphone camera is adopted to monitor in situ fluorescence signal from the reaction. Different from other existing CPCR systems with the described simple, easy-to-use, integrated, real-time microfluidic CPCR system, rapid nucleic acid analysis can be performed from sample to answer. A couple of critical issues, including wash scheme and reaction temperature, are analyzed for optimized system performance. It is demonstrated that influenza A virus with the reasonable concentration down to 1.0 TCID50/mL can be successfully detected by the integrated microfluidic system within 45 min.

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Section: Single Cpcr Reactor With Sample Preparationmentioning

confidence: 93%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

An Integrated, Real-Time Convective PCR System for Isolation, Amplification, and Detection of Nucleic Acids

Miao

Guo

et al. 2022

Chemosensors

Self Cite

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“…Nowadays, SP based PoCDs are also being explored as they have a simple optical attachment (camera) that can be used as a transducer for NAD. [ 37,38 ] This SP based flexible technology has been used to develop a cPCR amplification system using consumer‐class quadcopter drones technology “Lab‐on‐a‐drone” for rapid field deployment of NAD. [ 39,40 ] Simple, cost‐effective, and reusable platform based on composite polydimethylsiloxane (PDMS)‐on‐glass microchannel functionalized with peptide‐NA probes has been used to construct MFs streaming potential analyzer for NAD.…”

Section: Biomedical Diagnosticsmentioning

confidence: 99%

Emerging Trends in Microfluidics Based Devices

Solanki

Pandey

Gupta

et al. 2020

Biotechnology Journal

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One of the major challenges for scientists and engineers today is to develop technologies for the improvement of human health in both developed and developing countries. However, the need for cost-effective, high-performance diagnostic techniques is very crucial for providing accessible, affordable, and high-quality healthcare devices. In this context, microfluidic-based devices (MFDs) offer powerful platforms for automation and integration of complex tasks onto a single chip. The distinct advantage of MFDs lies in precise control of the sample quantities and flow rate of samples and reagents that enable quantification and detection of analytes with high resolution and sensitivity. With these excellent properties, microfluidics (MFs) have been used for various applications in healthcare, along with other biological and medical areas. This review focuses on the emerging demands of MFs in different fields such as biomedical diagnostics, environmental analysis, food and agriculture research, etc., in the last three or so years. It also aims to reveal new opportunities in these areas and future prospects of commercial MFDs.

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“…Based on PCR technology, heating media can be categorized into three types: water bath ( Chan et al, 2016 ), air heated ( Qiu et al, 2017 ), and metal heating block ( You et al, 2020 ). Water bath-based PCR has been abandoned because of its low degree of automation, low amplification efficiency, and lack of consistency ( Mahanama and Wilson-Davies, 2021 ).…”

Section: Introductionmentioning

confidence: 99%

Performance Evaluation of a Novel Ultrafast Molecular Diagnostic Device Integrated With Microfluidic Chips and Dual Temperature Modules

Lin

Song

Shao³

et al. 2022

Front. Bioeng. Biotechnol.

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Ultrafast, portable, and inexpensive molecular diagnostic platforms are critical for clinical diagnosis and on-site detection. There are currently no available real-time polymerase chain reaction (PCR) devices able to meet the demands of point-of-care testing, as the heating and cooling processes cannot be avoided. In this study, the dual temperature modules were first designed to process microfluidic chips automatically circulating between them. Thus, a novel ultrafast molecular diagnostic real-time PCR device (approximately 18 and 23 min for DNA and RNA detection, respectively) with two channels (FAM and Cy5) for the detection of 12 targets was developed. The device contained three core functional components, including temperature control, optics, and motion, which were integrated into a portable compact box. The temperature modules accurately control temperature in rapid thermal cycles with less than ±0.1 °C, ±1 °C and ±0.5 °C for the temperature fluctuation, uniformity, and error of indication, respectively. The average coefficient of variation (CV) of the fluorescence intensity (FI) for all 12 wells was 2.3% for FAM and 2.7% for Cy5. There was a good linear relationship between the concentrations of fluorescent dye and the FIs of FAM and Cy5(R2 = 0.9990 and 0.9937), and the average CVs of the Ct values calculated by the embedded software were 1.4% for FAM and Cy5, respectively. The 100 double-blind mocked sputum and 249 clinical stool samples were analyzed by the ultrafast real-time PCR device in comparison with the DAAN Gene SARS-CoV-2 kit run on the ABI 7500 instrument and Xpert C. difficile/Epi, respectively. Among the 249 stool samples, the ultrafast real-time PCR device detected toxigenic C. difficile in 54 samples (54/249, 21.7%) with a specificity and positive predictive values of 99.0 and 96.3%, which were higher than the Xpert C. difficile/Epi values of 94.4 and 88.1% (p > 0.05). The ultrafast real-time PCR device detected 15 SARS-CoV-2 positive samples, which has a 100% concordance with that obtained by the DAAN Gene SARS-CoV-2 kit. This study demonstrated that the ultrafast real-time PCR device integrated with microfluidic chips and dual temperature modules is an ultrafast, reliable, easy-to-use, and cost-effective molecular diagnostic platform for clinical diagnosis and on-site testing, especially in resource-limited settings.

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Characterization and analysis of real-time capillary convective PCR toward commercialization

Cited by 16 publications

References 40 publications

An Integrated, Real-Time Convective PCR System for Isolation, Amplification, and Detection of Nucleic Acids

An Integrated, Real-Time Convective PCR System for Isolation, Amplification, and Detection of Nucleic Acids

Emerging Trends in Microfluidics Based Devices

Performance Evaluation of a Novel Ultrafast Molecular Diagnostic Device Integrated With Microfluidic Chips and Dual Temperature Modules

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