scholarly journals Digital droplet PCR on disk

Lab on a Chip ◽  
2016 ◽  
Vol 16 (1) ◽  
pp. 208-216 ◽  
Author(s):  
Friedrich Schuler ◽  
Martin Trotter ◽  
Marcel Geltman ◽  
Frank Schwemmer ◽  
Simon Wadle ◽  
...  
Keyword(s):  
Pcr Amplification ◽  
Reaction Chamber ◽  
Droplet Generation ◽  
Digital Droplet Pcr ◽  
Droplet Pcr ◽  
Single Reaction

An integrated digital droplet PCR system is presented that enables droplet generation, PCR amplification, and readout within a single reaction chamber on a centrifugal microfluidic LabDisk.

scholarly journals The prevalence and diversity of AmpC β-lactamase genes in plasmids from aquatic systems

2018 ◽  
Vol 2017 (2) ◽  
pp. 603-611 ◽  
Author(s):  
Roelof Dirk Coertze ◽  
Cornelius Carlos Bezuidenhout
Keyword(s):  
South African ◽  
Aquatic Ecosystems ◽  
Antibiotic Resistance Genes ◽  
Pcr Amplification ◽  
Aquatic Systems ◽  
Bacterial Populations ◽  
Digital Droplet Pcr ◽  
Copy Numbers ◽  
Droplet Pcr ◽  
Plasmid Extraction

Abstract This study aimed to investigate the presence and diversity of AmpC β-lactamase and integrase genes among DNA (genomic and plasmid) from bacterial populations in selected aquatic systems. Following an enrichment step, DNA was isolated and subjected to polymerase chain reaction (PCR) and digital droplet PCR. The intI1 gene and AmpC β-lactamase genes were present in genomic and plasmid DNA from all sites in the Mooi, Crocodile and Marico Rivers, with the exception of intI1 in the Marico River. Digital droplet PCR demonstrated that copy numbers varied considerably (0.0 to 29.38 copies per picogram of DNA). Some samples in which ampC was not detected, intI1 was present. Amplicons of ampC genes were subjected to restriction digest using HindIII. Samples where the restriction markers were absent were purified by cloning followed by plasmid extraction, PCR amplification, and sequencing of individual AmpC gene fragments. Phylogenetic analysis identified all positive AmpC genes as Class C β-lactamases, comprising of ampC, CMY- and ACT-families. Detecting AmpC and intl1 genes on plasmids suggests a high risk of horizontal gene transfer and potential dissemination of these and other antibiotic resistance genes surrounding immediate aquatic environments. Consequences of β-lactamase diversity in aquatic ecosystems are relatively unexplored in South African aquatic ecosystems.

scholarly journals Monitoring EGFR C797S mutation in Japanese NSCLC patients with serial cell free DNA evaluation using digital droplet PCR

2021 ◽  
Author(s):  
Ryo Ariyasu ◽  
Ken Uchibori ◽  
Takaaki Sasaki ◽  
Mika Tsukahara ◽  
Kazuma Kiyotani ◽  
...  
Keyword(s):  
Cell Free Dna ◽  
Digital Droplet Pcr ◽  
Free Dna ◽  
Droplet Pcr ◽  
Nsclc Patients

scholarly journals P34.06 The Clinical Utility of Liquid Biopsy by Digital Droplet PCR in Patients with Advanced NSCLC

2021 ◽  
Vol 16 (3) ◽  
pp. S417
Author(s):  
V. Lamberts ◽  
M. Aldea ◽  
L. Mezquita ◽  
C. Jovelet ◽  
D. Vasseur ◽  
...  
Keyword(s):  
Liquid Biopsy ◽  
Clinical Utility ◽  
Advanced Nsclc ◽  
Digital Droplet Pcr ◽  
Droplet Pcr

Carbon metabolism snapshot by ddPCR during the early step of Candida albicans phagocytosis by macrophages

2020 ◽  
Vol 78 (1) ◽  
Author(s):  
Romain Laurian ◽  
Cécile Jacot-des-Combes ◽  
Fabiola Bastian ◽  
Karine Dementhon ◽  
Pascale Cotton
Keyword(s):  
Carbon Metabolism ◽  
Early Stage ◽  
Yeast Cells ◽  
Transcription Level ◽  
Digital Droplet Pcr ◽  
Metabolic Genes ◽  
Glucose Transport System ◽  
Short Period ◽  
Droplet Pcr ◽  
Alternative Sources

ABSTRACT During Candida macrophage interactions, phagocytosed yeast cells feed in order to grow, develop hyphae and escape. Through numerous proteomic and transcriptomic studies, two metabolic phases have been described. A shift to a starvation mode is generally identified as early as one-hour post phagocytosis, followed by a glycolytic growth mode after C. albicans escaped from the macrophage. Healthy macrophages contain low amounts of glucose. To determine if this carbon source was sensed and metabolized by the pathogen, we explored the transcription level of a delimited set of key genes expressed in C. albicans cells during phagocytosis by macrophages, at an early stage of the interaction. This analysis was performed using a technical digital droplet PCR approach to quantify reliably the expression of carbon metabolic genes after 30 min of phagocytosis. Our data confirm the technique of digital droplet PCR for the detection of C. albicans transcripts using cells recovered after a short period of phagocytosis. At this stage, carbon metabolism is clearly oriented towards the use of alternative sources. However, the activation of high-affinity glucose transport system suggests that the low amount of glucose initially present in the macrophages is detected by the pathogen.

scholarly journals Quantification of SARS-CoV-2 viral copy number in saliva mouthwash samples using digital droplet PCR

2020 ◽  
Author(s):  
Lisa Oberding ◽  
Jia Hu ◽  
Byron Berenger ◽  
Abu Naser Mohon ◽  
Dylan R. Pillai
Keyword(s):  
Viral Load ◽  
Copy Number ◽  
Clinical Samples ◽  
Digital Droplet Pcr ◽  
Viral Copy Number ◽  
Droplet Pcr ◽  
Precise Quantification ◽  
Viral Copy ◽  
Mouth Wash

AbstractSaliva samples were collected through a simple mouth wash procedure and viral load quantified using a technology called digital droplet PCR. Data suggest ddPCR allows for precise quantification of viral load in clinical samples infected with SARS-CoV-2.

Digital Droplet PCR (ddPCR) for Detection and Quantitation of Hepatitis Delt Virus

2021 ◽  
Author(s):  
Ling Xu ◽  
Xiangying Zhang ◽  
Yaling Cao ◽  
Zihao Fan ◽  
Yuan Tian ◽  
...  
Keyword(s):  
Liver Failure ◽  
Lower Limit ◽  
Limit Of Detection ◽  
Quantitative Detection ◽  
Rt Pcr ◽  
Rna Detection ◽  
Detection Rates ◽  
Digital Droplet Pcr ◽  
Droplet Pcr ◽  
Hdv Infection

Abstract Background & Aims The prevalence of hepatitis delt virus (HDV) far exceeds our expected level, there remains a lack of reliable quantitative assays for HDV RNA detection. We sought to develop a new method based on digital droplet PCR (ddPCR) for HDV RNA quantitative detection. Methods With plasmid (pMD19T) containing HDV full-genome, we determined the method for ddPCR-based HDV RNA quantification. To compare various assays for HDV detection, 30 cases diagnosed hepatitis D and 14 controls were examined by ELISA, RT-PCR and ddPCR. 728 HBV-related patients including 182 chronic hepatitis B (CHB), 182 liver cirrhosis (LC), 182 hepatocellular carcinoma (HCC) and 182 liver failure (LF) were screened for HDV infection. Results The limit of detection of ddPCR for HDV is significantly low, which lower limit of detection (LLoD) and lower limit of quantitation (LLoQ) to be 5.51 copies/reaction (95% CI: 1.15–6.4*105) and 0.18 copies/reaction (95% CI: 0.0012151- 0.76436), respectively. Among the 44 samples, ELISA detected 30 cases positive for anti-HDV, ddPCR reported 24 samples and RT-PCR reported 10 samples positive for HDV RNA. Moreover, the positive rates of anti-HDV IgG were 1.1%, 3.3%, 2.7% and 7.1% in patients with CHB, LC, HCC, and LF; the detection rates of RT-PCR in HDV RNA were 0%, 16.67%, 15.4% and 20%, however, the detection rates of ddPCR were 0%, 33.33%, 30.77% and 60%. Conclusion We establish a high sensitivity and high specificity quantitative HDV RNA detection method based on ddPCR compared to RT-PCR. HBV-related end-stage liver disease, especially liver failure, are associated with a remarkably high rate of HDV infection.

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