scholarly journals Detection of Copy Number Imbalance in Canine Urothelial Carcinoma With Droplet Digital Polymerase Chain Reaction

2015 ◽  
Vol 53 (4) ◽  
pp. 764-772 ◽  
Author(s):  
H. Mochizuki ◽  
S. G. Shapiro ◽  
M. Breen
Keyword(s):  
Polymerase Chain Reaction ◽  
Urothelial Carcinoma ◽  
Copy Number ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

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 Evaluation of a Droplet Digital Polymerase Chain Reaction Format for DNA Copy Number Quantification

2011 ◽  
Vol 84 (2) ◽  
pp. 1003-1011 ◽  
Author(s):  
Leonardo B. Pinheiro ◽  
Victoria A. Coleman ◽  
Christopher M. Hindson ◽  
Jan Herrmann ◽  
Benjamin J. Hindson ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Copy Number ◽  
Chain Reaction ◽  
Dna Copy Number ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

scholarly journals Validation of ctDNA Quality Control Materials Through a Precompetitive Collaboration of the Foundation for the National Institutes of Health

2021 ◽  
pp. 910-920
Author(s):  
P. Mickey Williams ◽  
Thomas Forbes ◽  
Steven P. Lund ◽  
Kenneth D. Cole ◽  
Hua-Jun He ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Quality Control ◽  
Copy Number ◽  
National Institutes Of Health ◽  
Single Nucleotide Variants ◽  
Chain Reaction ◽  
Single Nucleotide ◽  
Quality Control Materials ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

PURPOSE We report the results from a Foundation for the National Institutes of Health Biomarkers Consortium project to address the absence of well-validated quality control materials (QCMs) for circulating tumor DNA (ctDNA) testing. This absence is considered a cause of variance and inconsistencies in translating ctDNA results into clinical actions. METHODS In this phase I study, QCMs with 14 clinically relevant mutations representing single nucleotide variants, insertions or deletions (indels), translocations, and copy number variants were sourced from three commercial manufacturers with variant allele frequencies (VAFs) of 5%, 2.5%, 1%, 0.1%, and 0%. Four laboratories tested samples in quadruplicate using two allele-specific droplet digital polymerase chain reaction and three (amplicon and hybrid capture) next-generation sequencing (NGS) panels. RESULTS The two droplet digital polymerase chain reaction assays reported VAF values very close to the manufacturers’ claimed concentrations for all QCMs. NGS assays reported most single nucleotide variants and indels, but not translocations, close to the expected VAF values. Notably, two NGS assays reported lower VAF than expected for all translocations in all QCM mixtures, possibly related to technical challenges detecting these variants. The ability to call ERBB2 copy number amplifications varied across assays. All three QCMs provided valuable insight into assay precision. Each assay across all variant types demonstrated dropouts at 0.1%, suggesting that the QCM can serve for testing of an assay’s limit of detection with confidence claims for specific variants. CONCLUSION These results support the utility of the QCM in testing ctDNA assay analytical performance. However, unique designs and manufacturing methods for the QCM, and variations in a laboratory’s testing configuration, may require testing of multiple QCMs to find the best reagents for accurate result interpretation.

scholarly journals Digital Droplet PCR for SARS-CoV-2 Resolves Borderline Cases

2021 ◽  
Author(s):  
Jing Xu ◽  
Timothy Kirtek ◽  
Yan Xu ◽  
Hui Zheng ◽  
Huiyu Yao ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Limit Of Detection ◽  
False Negative ◽  
Accurate Method ◽  
Viral Envelope ◽  
Chain Reaction ◽  
Borderline Cases ◽  
Droplet Pcr ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Abstract Objectives The Bio-Rad SARS-CoV-2 ddPCR Kit (Bio-Rad Laboratories) was the first droplet digital polymerase chain reaction (ddPCR) assay to receive Food and Drug Administration (FDA) Emergency Use Authorization approval, but it has not been evaluated clinically. We describe the performance of ddPCR—in particular, its ability to confirm weak-positive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) results. Methods We clinically validated the Bio-Rad Triplex Probe ddPCR Assay. The limit of detection was determined by using serial dilutions of SARS-CoV-2 RNA in an artificial viral envelope. The ddPCR assay was performed according to the manufacturer’s specifications on specimens confirmed to be positive (n = 48) or negative (n = 30) by an FDA-validated reverse transcription–polymerase chain reaction assay on the m2000 RealTime system (Abbott). Ten borderline positive cases were also evaluated. Results The limit of detection was 50 copies/mL (19 of 20 positive). Forty-seven specimens spanning a range of quantification cycles (2.9-25.9 cycle numbers) were positive by this assay (47 of 48; 97.9% positive precent agreement), and 30 negative samples were confirmed as negative (30 of 30; 100% negative percent agreement). Nine of 10 borderline cases were positive when tested in triplicate. Conclusions The ddPCR of SARS-CoV-2 is an accurate method, with superior sensitivity for viral RNA detection. It could provide definitive evaluation of borderline positive cases or suspected false-negative cases.

Sign in / Sign up

Export Citation Format

Share Document