scholarly journals Utility of Real-Time Quantitative Polymerase Chain Reaction in DetectingMycobacterium tuberculosis

2017 ◽  
Vol 2017 ◽  
pp. 1-5 ◽  
Author(s):  
Zhongquan Lv ◽  
Mingxin Zhang ◽  
Hui Zhang ◽  
Xinxin Lu
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Roc Curve ◽  
Quantitative Polymerase Chain Reaction ◽  
Culture Method ◽  
Gene Copy ◽  
Youden Index ◽  
Chain Reaction ◽  
Cutoff Value ◽  
Polymerase Chain

This study aimed to assess the value of real-time quantitative polymerase chain reaction (RT-qPCR) for the detection ofMycobacterium tuberculosis(MTB). Samples from 192 patients with suspected MTB were examined by RT-qPCR and an improved Löwenstein–Jensen (L-J) culture method. To evaluate the diagnostic usefulness of RT-qPCR in detecting MTB, a receiver operating characteristic (ROC) curve for RT-qPCR was generated, and the area under the curve (AUC) as well as a cutoff value was calculated. Using the L-J culture method as the gold standard, accuracy of the RT-qPCR method for detecting MTB was 92.7%, with sensitivity and specificity of 62.5% and 97.02%, respectively. In comparison with the improved L-J culture method, the AUC of RT-qPCR ROC curve was 0.957, which was statistically significant (p<0.001). The Youden Index reached the maximum value (0.88) for gene copy number of 794.5 IU/mL, which was used as the cutoff value. RT-qPCR detection of MTB yielded results consistent with those of the improved L-J culture method, with high accuracy. RT-qPCR may be used as an auxiliary method for etiological diagnosis of tuberculosis.

scholarly journals Brief communication (Original). Rapid diagnosis of trisomy 21 by relative gene copy using real-time quantitative polymerase chain reaction

2014 ◽  
Vol 8 (3) ◽  
pp. 399-403
Author(s):  
Chinnuwat Sanguansermsri ◽  
Pranoot Tanpaiboon ◽  
Pimlak Charoenkwan ◽  
Arunee Phusua
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Trisomy 21 ◽  
Diagnostic Method ◽  
Quantitative Polymerase Chain Reaction ◽  
Chromosome 21 ◽  
Gene Copy ◽  
Chain Reaction ◽  
Karyotypic Analysis ◽  
Polymerase Chain

Abstract Background: Trisomy 21 or Down syndrome (DS) is the most common aneuploidy disorder. Fetal karyotypic analysis remains the criterion standard for prenatal diagnosis of DS, although the method is time consuming and requires skilled personnel. Real-time quantitative polymerase chain reaction (qPCR) can be used to determine a difference in the amount of gene copy by calculation of the difference between the cycle threshold (ΔCT) of a tested gene and a reference gene. Objectives: To develop a rapid qPCR diagnostic method for trisomy 21. Methods: Ten DS patients with the known karyotype of trisomy 21 were enrolled. Their parents were included as controls. D21S11 locus on chromosome 21 and SM locus on chromosome 16 from each subject were amplified by qPCR. The D21S11/SM ΔCT and 2-ΔΔCT values were compared between DS patients and their parents. Results: The D21S11/SM ΔCT values of the DS patients were higher than their respective controls except for one family. The mean 2-ΔΔCT value between patients and mothers was 1.88 ± 0.95 (95% CI 1.20-2.56), and between fathers and mothers as controls was 1.06 ± 0.68 (95% CI 0.58-1.54). Conclusion: The diagnostic method of trisomy 21 by using qPCR is feasible, although false negative results may occur. Using more index genes is recommended to increase the sensitivity and specificity.

scholarly journals Analysis and validation of a highly sensitive one-step nested quantitative real-time polymerase chain reaction assay for specific detection of severe acute respiratory syndrome coronavirus 2

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Yang Zhang ◽  
Chunyang Dai ◽  
Huiyan Wang ◽  
Yong Gao ◽  
Tuantuan Li ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Quantitative Polymerase Chain Reaction ◽  
Clinical Samples ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Qrt Pcr ◽  
Highly Sensitive ◽  
One Step ◽  
Polymerase Chain

Abstract Background Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, is posing a serious threat to global public health. Reverse transcriptase real-time quantitative polymerase chain reaction (qRT-PCR) is widely used as the gold standard for clinical detection of SARS-CoV-2. Due to technical limitations, the reported positive rates of qRT-PCR assay of throat swab samples vary from 30 to 60%. Therefore, the evaluation of alternative strategies to overcome the limitations of qRT-PCR is required. A previous study reported that one-step nested (OSN)-qRT-PCR revealed better suitability for detecting SARS-CoV-2. However, information on the analytical performance of OSN-qRT-PCR is insufficient. Method In this study, we aimed to analyze OSN-qRT-PCR by comparing it with droplet digital PCR (ddPCR) and qRT-PCR by using a dilution series of SARS-CoV-2 pseudoviral RNA and a quality assessment panel. The clinical performance of OSN-qRT-PCR was also validated and compared with ddPCR and qRT-PCR using specimens from COVID-19 patients. Result The limit of detection (copies/ml) of qRT-PCR, ddPCR, and OSN-qRT-PCR were 520.1 (95% CI: 363.23–1145.69) for ORF1ab and 528.1 (95% CI: 347.7–1248.7) for N, 401.8 (95% CI: 284.8–938.3) for ORF1ab and 336.8 (95% CI: 244.6–792.5) for N, and 194.74 (95% CI: 139.7–430.9) for ORF1ab and 189.1 (95% CI: 130.9–433.9) for N, respectively. Of the 34 clinical samples from COVID-19 patients, the positive rates of OSN-qRT-PCR, ddPCR, and qRT-PCR were 82.35% (28/34), 67.65% (23/34), and 58.82% (20/34), respectively. Conclusion In conclusion, the highly sensitive and specific OSN-qRT-PCR assay is superior to ddPCR and qRT-PCR assays, showing great potential as a technique for detection of SARS-CoV-2 in patients with low viral loads.

Quantification of Common Wheat Adulteration of Durum Wheat Pasta Using Real-Time Quantitative Polymerase Chain Reaction (PCR)

2002 ◽  
Vol 79 (4) ◽  
pp. 553-558 ◽  
Author(s):  
Rémi Alary ◽  
Arnaud Serin ◽  
Marie-Pierre Duviau ◽  
Philippe Jourdrier ◽  
Marie-Françoise Gautier
Keyword(s):  
Polymerase Chain Reaction ◽  
Durum Wheat ◽  
Real Time ◽  
Common Wheat ◽  
Quantitative Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Polymerase Chain

Quantitative Real-Time Polymerase Chain Reaction Detection of BK Virus Using Labeled Primers

2010 ◽  
Vol 134 (3) ◽  
pp. 444-448 ◽  
Author(s):  
Zhengming Gu ◽  
Jianmin Pan ◽  
Matthew J. Bankowski ◽  
Randall T. Hayden
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Real Time Pcr ◽  
Quantitative Polymerase Chain Reaction ◽  
Bk Virus ◽  
Clinical Samples ◽  
Quantitative Detection ◽  
Chain Reaction ◽  
Pcr Method ◽  
Polymerase Chain

Abstract Context.—BK virus infections among immunocompromised patients are associated with disease of the kidney or urinary bladder. High viral loads, determined by quantitative polymerase chain reaction (PCR), have been correlated with clinical disease. Objective.—To develop and evaluate a novel method for real-time PCR detection and quantification of BK virus using labeled primers. Design.—Patient specimens (n = 54) included 17 plasma, 12 whole blood, and 25 urine samples. DNA was extracted using the MagNA Pure LC Total Nucleic Acid Isolation Kit (Roche Applied Science, Indianapolis, Indiana); sample eluate was PCR-amplified using the labeled primer PCR method. Results were compared with those of a user-developed quantitative real-time PCR method (fluorescence resonance energy transfer probe hybridization). Results.—Labeled primer PCR detected less than 10 copies per reaction and showed quantitative linearity from 101 to 107 copies per reaction. Analytical specificity of labeled primer PCR was 100%. With clinical samples, labeled primer PCR demonstrated a trend toward improved sensitivity compared with the reference method. Quantitative assay comparison showed an R2 value of 0.96 between the 2 assays. Conclusions.—Real-time PCR using labeled primers is highly sensitive and specific for the quantitative detection of BK virus from a variety of clinical specimens. These data demonstrate the applicability of labeled primer PCR for quantitative viral detection and offer a simplified method that removes the need for separate oligonucleotide probes.

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