scholarly journals Detection and quantification of chimerism by droplet digital PCR

Chimerism ◽  
2013 ◽  
Vol 4 (3) ◽  
pp. 102-108 ◽  
Author(s):  
David George ◽  
Juliann Czech ◽  
Bobby John ◽  
Min Yu ◽  
Lawrence J. Jennings
Keyword(s):  
Digital Pcr ◽  
Droplet Digital Pcr ◽  
Detection And Quantification

scholarly journals Development and Evaluation of a Single Dye Duplex Droplet Digital PCR Assay for the Rapid Detection and Quantification of Mycobacterium tuberculosis

2020 ◽  
Vol 8 (5) ◽  
pp. 701 ◽  
Author(s):  
Raphael Nyaruaba ◽  
Jin Xiong ◽  
Caroline Mwaliko ◽  
Nuo Wang ◽  
Belindah J. Kibii ◽  
...  
Keyword(s):  
Target Genes ◽  
Annealing Temperature ◽  
Single Channel ◽  
Digital Pcr ◽  
Absolute Quantification ◽  
Droplet Digital Pcr ◽  
Sample Concentration ◽  
Specificity And Sensitivity ◽  
Probe Concentration ◽  
Detection And Quantification

Droplet digital PCR (ddPCR) is a third generation of PCR that was recently developed to overcome the challenges of real-time fluorescence-based quantitative PCR (qPCR) in absolute quantification of pathogens. Few studies have been done on tuberculosis (TB) detection and quantification using ddPCR despite its many advantages over qPCR. From the few studies, none explores a single dye duplex assay for the detection and quantification of TB. In this study, steps toward developing and evaluating a duplex single dye (FAM) assay for detecting two targets (IS6110 and IS1081) are clearly described using simplex and duplex experiments. To achieve this, various parameters are investigated, including annealing temperature, primer and probe concentration, sensitivity and specificity, sample concentration, and inter/intra-assay variability. From the results, primer and probe concentration, annealing temperature, and sample concentration have an effect on the position and separation of droplets in both simplex and duplex assays. The copies of target genes in a duplex assay can be estimated accurately using the threshold tool with little inter-assay (CV <1%) and intra-assay (CV <6%) variability when compared to simplex assays. The ddPCR assay specificity and sensitivity are both 100% when compared to qPCR. This work shows steps toward the detection and quantification of two targets in a single channel, enabling higher multiplexing to include more targets in future works.

scholarly journals Droplet digital PCR for detection and quantification of circulating tumor DNA in plasma of head and neck cancer patients

BMC Cancer ◽  
2017 ◽  
Vol 17 (1) ◽  
Author(s):  
Joost H. van Ginkel ◽  
Manon M. H. Huibers ◽  
Robert J. J. van Es ◽  
Remco de Bree ◽  
Stefan M. Willems
Keyword(s):  
Head And Neck Cancer ◽  
Head And Neck ◽  
Cancer Patients ◽  
Neck Cancer ◽  
Digital Pcr ◽  
Circulating Tumor Dna ◽  
Droplet Digital Pcr ◽  
Tumor Dna ◽  
Detection And Quantification

scholarly journals Effect of Amount of DNA on Digital PCR Assessment of Genetically Engineered Canola and Soybean Events

2020 ◽  
Author(s):  
Tigst Demeke ◽  
Monika Eng ◽  
Michelle Holigroski ◽  
Sung-Jong Lee
Keyword(s):  
Zero Tolerance ◽  
Digital Pcr ◽  
Absolute Quantification ◽  
Genetically Engineered ◽  
Droplet Digital Pcr ◽  
Chain Reaction ◽  
Low Level ◽  
Polymerase Chain ◽  
Reliable Quantification ◽  
Detection And Quantification

Abstract Low-level detection and quantification of genetically engineered (GE) traits with polymerase chain reaction (PCR) is challenging. For unapproved GE events, any level of detection is not acceptable in some countries because of zero tolerance. Droplet digital PCR (ddPCR) has been successfully used for absolute quantification of GE events. In this study, reliability of low level quantification of GE events with ddPCR was assessed using a total of 50, 100, 200, 400, and 600 ng DNA spiked at 0.01% and 0.1% concentration levels. Genetically engineered canola (GT73 and MON88302 events) and soybean (A2704-12 and DP305423 events) events were used for the study. For samples spiked at 0.1% level, reliable quantification was achieved for the four GE events using 50 or 100 ng DNA. Few target droplets were generated for 0.01% spiked GE samples using 50 and 100 ng DNA. Increasing the amount of DNA for ddPCR generated more number of target droplets. For GE canola events, the use of 400 and 600 ng DNA for ddPCR resulted in saturation. The use of multiple wells of 200 ng DNA (instead of 400 and 600 ng per well) helped to overcome the saturation problem. Overall, the use of high amount of DNA for ddPCR was helpful for the detection and quantification of 0.01% GE samples.

Two-Step Reverse Transcription Droplet Digital PCR Protocols for SARS-CoV-2 Detection and Quantification

10.3791/62295 ◽  
2021 ◽  
Author(s):  
Raphael Nyaruaba ◽  
Xiaohong Li ◽  
Caroline Mwaliko ◽  
Changchang Li ◽  
Matilu Mwau ◽  
...  
Keyword(s):  
Reverse Transcription ◽  
Digital Pcr ◽  
Droplet Digital Pcr ◽  
Detection And Quantification

scholarly journals Detection and Quantification of BCR-ABL1 Fusion Transcripts by Droplet Digital PCR

2014 ◽  
Vol 16 (2) ◽  
pp. 174-179 ◽  
Author(s):  
Lawrence J. Jennings ◽  
David George ◽  
Juliann Czech ◽  
Min Yu ◽  
Loren Joseph
Keyword(s):  
Digital Pcr ◽  
Droplet Digital Pcr ◽  
Fusion Transcripts ◽  
Detection And Quantification
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