Direct capture and amplification of nucleic acids using a universal, elution-free magnetic bead-based method for rapid pathogen detection in multiple types of biological samples

Jiang, Qianqian; Li, Yang; Huang, Lin; Guo, Jun; Wang, Ailin; Ma, Cuiping; Shi, Chao

doi:10.1007/s00216-022-04422-8

Cited by 4 publications

(6 citation statements)

References 32 publications

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“…First, as our device currently only detects HIV RNA, sample preparation (e.g., HIV RNA extraction) still needs to be incorporated. To this end, nucleic acid extraction based on magnetic beads (Fan et al, 2023;Jiang et al, 2023;Kang et al, 2021;Ngo et al, 2023;Pearlman et al, 2020;Trick et al, 2022), cellulose paper (Zou et al, 2017), or FTA membrane (Kadimisetty et al, 2021;Ndunguru et al, 2005) offer possible solutions. It may also be feasible to adopt a custom digital chip with integrated nucleic acid extraction capability (Yin et al, 2020).…”

Section: Discussionmentioning

confidence: 99%

Rapid and Portable Quantification of HIV RNA via a Smartphone-enabled Digital CRISPR Device and Deep Learning

Ngo

Akarapipad

Lee

et al. 2023

Preprint

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For the 28.2 million people in the world living with HIV/AIDS and receiving antiretroviral therapy, it is crucial to monitor their HIV viral loads with ease. To this end, rapid and portable diagnostic tools that can quantify HIV RNA are critically needed. We report herein a rapid and quantitative digital CRISPR-assisted HIV RNA detection assay that has been implemented within a portable smartphone-based device as a potential solution. Specifically, we first developed a fluorescence-based reverse transcription recombinase polymerase amplification (RT-RPA)-CRISPR assay for isothermally and rapidly detecting HIV RNA at 42 C in < 30 min. When realized within a commercial stamp-sized digital chip, this assay yields strongly fluorescent digital reaction wells corresponding to HIV RNA. The isothermal reaction condition and the strong fluorescence in the small digital chip unlock compact thermal and optical components in our device, allowing us to engineer a palm-size (70 by 115 by 80 mm) and lightweight (< 0.6 kg) device. Further leveraging the smartphone, we wrote a custom app to control the device, perform the digital assay, and acquire fluorescence images throughout the assay time. We additionally trained and verified a Deep Learning-based algorithm for analyzing fluorescence images and detecting strongly fluorescent digital reaction wells. Using our smartphone-enabled digital CRISPR device, we were able to detect 75 copies of HIV RNA in 15 min and demonstrate the potential of our device toward convenient monitoring of HIV viral loads and combating the HIV/AIDS epidemic.

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Section: Discussionmentioning

confidence: 99%

Rapid and Portable Quantification of HIV RNA via a Smartphone-enabled Digital CRISPR Device and Deep Learning

Ngo

Akarapipad

Lee

et al. 2023

Preprint

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“…Staphylococcus aureus (ATCC 25923) and Escherichia coli O157:H7 (ATCC 35150) strains were provided by JianMa Gene Technology Co., Ltd. (Qingdao, China). PCR and LAMP reaction mixtures were prepared according to our previous work . A bacterial ROS Assay Kit was purchased from Jining Industrial Co., Ltd. (Shanghai, China).…”

Section: Experimental Sectionmentioning

confidence: 99%

“…All amplification reactions were performed using a CFX Connect Real-Time PCR System (Bio-Rad, CA), as described previously. 28 Assessment of Oxidative Stress Response Triggered by DFs. The oxidative stress response triggered by DFs was evaluated by measuring the changes in ROS and relative gene expression levels in S. aureus and E. coli O157:H7 cells.…”

Section: ■ Introductionmentioning

confidence: 99%

Lysis-Free Isolation and Direct Amplification of Pathogenic Bacterial DNA Using Diatom Frustules

Li,

Sun,

Wang

et al. 2024

Anal. Chem.

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DNA has been implicated as an important biomarker for the diagnosis of bacterial infections. Herein, we developed a streamlined methodology that uses diatom frustules (DFs) to liberate and capture bacterial DNA and allows direct downstream amplification tests without any lysis, washing, or elution steps. Unlike most conventional DNA isolation methods that rely on cell lysis to release bacterial DNA, DFs can trigger the oxidative stress response of bacterial cells to promote bacterial membrane vesicle formation and DNA release by generating reactive oxygen species in aqueous solutions. Due to the hierarchical porous structure, DFs provided high DNA capture efficiency exceeding 80% over a wide range of DNA amounts from 10 pg to 10 ng, making only 10 μg DFs sufficient for each test. Since laborious liquid handling steps are not required, the entire DNA sample preparation process using DFs can be completed within 3 min. The diagnostic use of this DF-based methodology was illustrated, which showed that the DNA of the pathogenic bacteria in serum samples was isolated by DFs and directly detected using polymerase chain reaction (PCR) at concentrations as low as 102 CFU/mL, outperforming the most used approaches based on solid-phase DNA extraction. Furthermore, most of the bacterial cells were still alive after DNA isolation using DFs, providing the possibility of recycling samples for storage and further diagnosis. The proposed DF-based methodology is anticipated to simplify bacterial infection diagnosis and be broadly applied to various medical diagnoses and biological research.

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“…With rapid developments in the field of molecular biology, precision diagnosis, and treatment, the demand for emerging nucleic acid extraction technologies with high throughput, purity, and quality is constantly increasing. As a simple, fast, reliable and automated nucleic acid extraction method, the magnetic beads(MBs) method [70] for nucleic acid extraction has attracted more and more attention. The process of DNA extraction by the MBs method is simple, without repeated centrifugation, column separation, or vacuum filtration [71].…”

Section: Nucleic Acid Extraction Technologymentioning

confidence: 99%

“…This solution solves the drawbacks of magnetic bead-based solid-phase extraction that may cause nucleic acid loss due to the handling of magnetic beads when they are transferred from one chamber to another, resulting in higher yield and purity in the end. Jiang et al [70] and others established a no-elution MB-based nucleic acid extraction method by introducing PEPPG F68 into the lysate and using NaOH solution instead of alcohol as the washing buffer. It avoids the dilution and loss of the target nucleic acid during the elution process, as well as the possible loss of sensitivity and false-negative results.…”

Section: Nucleic Acid Extraction Technologymentioning

confidence: 99%

Research Progress of Nucleic Acid Detection Technology for Genetically Modified Maize

Luo,

Li,

Wang

et al. 2023

IJMS

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Genetically modified (GM) maize is one of the earliest GM crops to have achieved large-scale commercial cultivation globally, and it is of great significance to excel in the development and implementation of safety policy regarding GM, and in its technical oversight. This article describes the general situation regarding genetically modified maize, including its varieties, applications, relevant laws and regulations, and so on. From a technical point of view, we summarize and critically analyze the existing methods for detecting nucleic acid levels in genetically modified maize. The nucleic acid extraction technology used for maize is explained, and the introduction of traditional detection techniques, which cover variable-temperature and isothermal amplification detection technology and gene chip technology, applications in maize are described. Moreover, new technologies are proposed, with special attention paid to nucleic acid detection methods using sensors. Finally, we review the current limitations and challenges of GM maize nucleic acid testing and share our vision for the future direction of this field.

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Direct capture and amplification of nucleic acids using a universal, elution-free magnetic bead-based method for rapid pathogen detection in multiple types of biological samples

Cited by 4 publications

References 32 publications

Rapid and Portable Quantification of HIV RNA via a Smartphone-enabled Digital CRISPR Device and Deep Learning

Rapid and Portable Quantification of HIV RNA via a Smartphone-enabled Digital CRISPR Device and Deep Learning

Lysis-Free Isolation and Direct Amplification of Pathogenic Bacterial DNA Using Diatom Frustules

Research Progress of Nucleic Acid Detection Technology for Genetically Modified Maize

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