scholarly journals Ultrasensitive Detection of Pb2+ Based on a DNAzyme and Digital PCR

2019 ◽  
Vol 2019 ◽  
pp. 1-6 ◽  
Author(s):  
Tao Zhang ◽  
Cong Liu ◽  
Wuping Zhou ◽  
Keming Jiang ◽  
Chenyu Yin ◽  
...  
Keyword(s):  
Detection Method ◽  
Detection System ◽  
Digital Pcr ◽  
Quantitative Detection ◽  
Ultrasensitive Detection ◽  
Hydrolytic Cleavage ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Template Dna ◽  
Digital Polymerase Chain Reaction

In this study, an ultrasensitive detection method for aqueous Pb2+ based on digital polymerase chain reaction (dPCR) technology and a Pb2+-dependent DNAzyme was developed. In the presence of Pb2+, the Gr-5 DNAzyme was activated and catalyzed the hydrolytic cleavage of the substrate strand, resulting in an increase in the amount of template DNA available for dPCR and a resultant change in the number of droplets showing a positive signal. Moreover, the detection system was found to be sensitive and stable in environmental sample detection. In summary, an ultrasensitive quantitative detection method for Pb2+ within environmental substrates was established.

scholarly journals Droplet Digital PCR (ddPCR) Analysis for the Detection and Quantification of Cow DNA in Buffalo Mozzarella Cheese

Animals ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1270
Author(s):  
Anna Cutarelli ◽  
Andrea Fulgione ◽  
Pasquale Fraulo ◽  
Francesco Paolo Serpe ◽  
Pasquale Gallo ◽  
...  
Keyword(s):  
Digital Pcr ◽  
Buffalo Milk ◽  
Cow Milk ◽  
Mozzarella Cheese ◽  
Chain Reaction ◽  
New Methods ◽  
Protected Designation Of Origin ◽  
Commission Regulation ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Buffalo mozzarella cheese is one of the most appreciated traditional Italian products and it is certified as a Protected Designation of Origin (PDO) product under the European Commission Regulation No. 1151/2012. It is obtained exclusively from buffalo milk. If made from cow milk, or a mixture of buffalo and cow milk, buffalo mozzarella cheese does not qualify as a PDO product. In order to maximize their profits, some producers market buffalo mozzarella that also contains cow milk as a PDO product, thus defrauding consumers. New methods for revealing this fraud are therefore needed. One such method is the droplet digital Polymerase Chain Reaction (ddPCR). Thanks to its high precision and sensitivity, the ddPCR could prove an efficacious means for detecting the presence of cow milk in buffalo mozzarella cheese that is marketed as a PDO product. ddPCR has proved able to detect the DNA of cow and/or buffalo milk in 33 buffalo mozzarella cheeses labelled as PDO products, and experimental evidence could support its application in routine analyses.

Quantitative detection of human enteric adenoviruses in river water by microfluidic digital polymerase chain reaction

2014 ◽  
Vol 70 (3) ◽  
pp. 555-560 ◽  
Author(s):  
Naohiro Kishida ◽  
Naohiro Noda ◽  
Eiji Haramoto ◽  
Mamoru Kawaharasaki ◽  
Michihiro Akiba ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
River Water ◽  
Water Samples ◽  
Real Time Pcr ◽  
Quantitative Detection ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
Enteric Adenoviruses

We describe an assay for simple and accurate quantification of human enteric adenoviruses (EAdVs) in water samples using a recently developed quantification method named microfluidic digital polymerase chain reaction (dPCR). The assay is based on automatic distribution of reaction mixture into a large number of nanolitre-volume reaction chambers and absolute copy number quantification from the number of chambers containing amplification products on the basis of Poisson statistics. This assay allows absolute quantification of target genes without the use of standard DNA. Concentrations of EAdVs in Japanese river water samples were successfully quantified by the developed dPCR assay. The EAdVs were detected in seven of the 10 samples (1 L each), and the concentration ranged from 420 to 2,700 copies/L. The quantified values closely resemble those by most probable number (MPN)-PCR and real-time PCR when standard DNA was validated by dPCR whereas they varied substantially when the standard was not validated. Accuracy and sensitivity of the dPCR was higher than those of real-time PCR and MPN-PCR. To our knowledge, this is the first study that has successfully quantified enteric viruses in river water using dPCR. This method will contribute to better understanding of existence of viruses in water.

scholarly journals Quantitative Detection of Beef and Beef Meat Products Adulteration by the Addition of Duck Meat Using Micro Drop Digital Polymerase Chain Reaction

2020 ◽  
Vol 2020 ◽  
pp. 1-8
Author(s):  
Chen Chen ◽  
Jia Chen ◽  
Yan Zhang ◽  
Yongbo Li ◽  
Zan Wang ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Analysis ◽  
Copy Number ◽  
Meat Products ◽  
Quantitative Detection ◽  
Chain Reaction ◽  
Dna Copy Number ◽  
Meat Weight ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

A single-copy specific primer was designed based on beef and duck samples and through drop digital polymerase chain reaction (ddPCR) for the quantitative analysis. Results revealed that the primers had no specific amplification with sheep, chicken, pork, or other species. Both the relationships between meat weight and DNA weight and between DNA weight and DNA copy number (C) were nearly linear within the dynamic range. To calculate the original meat weight from the DNA copy number, the DNA weight was used as the intermediate value to establish the following formulae: Mbeef = 0.058C − 1.86; Mduck = 0.0268C − 7.78. To achieve a good quantitative analysis, all species used in the experiment were made of lean meat. The accuracy of the method was verified by artificial adulteration of different proportions. Testing of the commercial samples indicated that adulteration is present in the market. The established digital PCR method provided an effective tool for monitoring the adulterated meat products and reducing the adulteration in the market.

scholarly journals Detection of fish allergen by droplet digital PCR

2019 ◽  
Vol 7 (4) ◽  
Author(s):  
Cinzia Daga ◽  
Simona Cau ◽  
Maria Giovanna Tilocca ◽  
Barbara Soro ◽  
Aldo Marongiu ◽  
...  
Keyword(s):  
18S Rrna ◽  
Annealing Temperature ◽  
Pcr Primers ◽  
Digital Pcr ◽  
18S Rrna Gene ◽  
Rrna Gene ◽  
Chain Reaction ◽  
European Regulation ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Fish is one of fourteen allergens that must be highlighted on the label within the ingredients list. It should be noted that the European regulation, is very restrictive to allergens with zero tolerance. Therefore it is important to establish sensitive and specific methods for detecting fish allergen. Applicability to detect and quantify fish allergen by droplet digital polymerase chain reaction (ddPCR) has been evaluated in this work. Genomic DNA of three fish species belonging to the most common fish families were analyzed. PCR primers were designed to amplify a 166 bp region of the 18S rRNA gene. Comparative studies were performed to establish the optimal primer and probe concentrations.  Annealing temperature was determined by using thermal gradient. The results have shown good applicability of the optimized 18S rRNA gene-method to detect and quantify small amounts of the target in all samples analyzed. However, validation studies are needed in order to apply ddPCR technology for routine allergens analysis.  

scholarly journals Digital PCR: A Reliable Tool for Analyzing and Monitoring Hematologic Malignancies

2020 ◽  
Vol 21 (9) ◽  
pp. 3141 ◽  
Author(s):  
Nicoletta Coccaro ◽  
Giuseppina Tota ◽  
Luisa Anelli ◽  
Antonella Zagaria ◽  
Giorgina Specchia ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Residual Disease ◽  
Hematologic Malignancies ◽  
Digital Pcr ◽  
Future Perspectives ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
Generation Sequencing ◽  
Minimal Residual

The digital polymerase chain reaction (dPCR) is considered to be the third-generation polymerase chain reaction (PCR), as it yields direct, absolute and precise measures of target sequences. dPCR has proven particularly useful for the accurate detection and quantification of low-abundance nucleic acids, highlighting its advantages in cancer diagnosis and in predicting recurrence and monitoring minimal residual disease, mostly coupled with next generation sequencing. In the last few years, a series of studies have employed dPCR for the analysis of hematologic malignancies. In this review, we will summarize these findings, attempting to focus on the potential future perspectives of the application of this promising technology.

scholarly journals Application of droplet digital polymerase chain reaction of plasma methylated septin 9 on detection and early monitoring of colorectal cancer

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Zhi Yao Ma ◽  
Cherry Sze Yan Chan ◽  
Kam Shing Lau ◽  
Lui Ng ◽  
Yuen Yee Cheng ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Polymerase Chain Reaction ◽  
Area Under The Curve ◽  
Quantitative Detection ◽  
Chain Reaction ◽  
Septin 9 ◽  
Non Invasive ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

AbstractMethylated septin 9 (SEPT9) has been approved for non-invasive screening of colorectal cancer (CRC), but data on monitoring of CRC is sparse. Droplet digital polymerase chain reaction (ddPCR), with higher detection precision and simpler quantification than conventional PCR, has not been applied in SEPT9 detection. We explored the role of SEPT9 ddPCR for CRC detection and to measure serial SEPT9 levels in blood samples of CRC patients before and 3-month after surgery. SEPT9 methylated ratio, methylated abundance, and CEA levels were all higher in CRC patients than normal controls (all P < 0.05). The area under the curve (AUC) for methylated ratio and abundance to detect CRC was 0.707 and 0.710, respectively. There was an increasing trend for SEPT9 methylated abundance from proximal to distal cancers (P = 0.017). At 3-month after surgery, both methylated abundance and ratio decreased (P = 0.005 and 0.053, respectively), especially methylated abundance in stage III and distal cancer (both P < 0.01). We have developed a ddPCR platform for the quantitative detection of plasma SEPT9 in CRC patients. SEPT9 methylated abundance had an early post-operative decline, which may be useful in monitoring of treatment response.

Heterogeneous modification of through-hole microwell chips for ultralow cross-contamination digital polymerase chain reaction

The Analyst ◽  
2020 ◽  
Vol 145 (8) ◽  
pp. 3116-3124
Author(s):  
Jinze Li ◽  
Yajun Qiu ◽  
Zhiqi Zhang ◽  
Chuanyu Li ◽  
Shuli Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Digital Pcr ◽  
Cross Contamination ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
Through Hole

Heterogeneous modification of through-hole microwell chips to avoid cross-contamination during digital PCR.

scholarly journals Application of droplet digital PCR to detect the pathogens of infectious diseases

2018 ◽  
Vol 38 (6) ◽  
Author(s):  
Haiyi Li ◽  
Ruolan Bai ◽  
Zhenyu Zhao ◽  
Lvyan Tao ◽  
Mingbiao Ma ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Infectious Diseases ◽  
Quantitative Polymerase Chain Reaction ◽  
Digital Pcr ◽  
Conventional Polymerase Chain Reaction ◽  
Nucleic Acid Detection ◽  
Chain Reaction ◽  
Molecular Biology Technique ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Polymerase chain reaction (PCR) is a molecular biology technique used to multiply certain deoxyribonucleic acid (DNA) fragments. It is a common and indispensable technique that has been applied in many areas, especially in clinical laboratories. The third generation of polymerase chain reaction, droplet digital polymerase chain reaction (ddPCR), is a biotechnological refinement of conventional polymerase chain reaction methods that can be used to directly quantify and clonally amplify DNA. Droplet digital polymerase chain reaction is now widely used in low-abundance nucleic acid detection and is useful in diagnosis of infectious diseases. Here, we summarized the potential advantages of droplet digital polymerase chain reaction in clinical diagnosis of infectious diseases, including viral diseases, bacterial diseases and parasite infections, concluded that ddPCR provides a more sensitive, accurate, and reproducible detection of low-abundance pathogens and may be a better choice than quantitative polymerase chain reaction for clinical applications in the future.

A rapid nucleic acid concentration measurement system with large-field-of-view for droplet digital PCR microfluidic chip

Lab on a Chip ◽  
2021 ◽  
Author(s):  
Jinrong Shen ◽  
Jihong Zheng ◽  
Zhenqing Li ◽  
Yourong Liu ◽  
Fengxiang Jing ◽  
...  
Keyword(s):  
Digital Pcr ◽  
Absolute Quantification ◽  
Droplet Digital Pcr ◽  
Field Of View ◽  
Large Field ◽  
Chain Reaction ◽  
Effective Technique ◽  
Nucleic Acid Concentration ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Droplet digital polymerase chain reaction(ddPCR) is an effective technique for the absolute quantification of target mucleic acid unparalleled sensitivity. However, current commerical ddPCR device for the detection of the gene...

A localized temporary negative pressure assisted microfluidic device for detecting keratin 19 in A549 lung carcinoma cells with digital PCR

2015 ◽  
Vol 7 (5) ◽  
pp. 2006-2011 ◽  
Author(s):  
Qingchang Tian ◽  
Qi Song ◽  
Yanan Xu ◽  
Qiangyuan Zhu ◽  
Bingwen Yu ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Digital Pcr ◽  
Carcinoma Cells ◽  
Biological Research ◽  
Chain Reaction ◽  
Lung Carcinoma Cells ◽  
Keratin 19 ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction ◽  
A549 Lung

Digital polymerase chain reaction (dPCR) has played a major role in biological research, especially by providing an accurate counting of single nucleic acid molecules.

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