scholarly journals Development of plasmids for quantitative detection of integrated lentiviral vectors and evaluation of culture time to perform vector titer by real-time quantitative polymerase chain reaction assay

2014 ◽  
Vol 4 (2) ◽  
Author(s):  
Elena Baiamonte ◽  
Mariella Bagliesi ◽  
Valentina Motta ◽  
Barbara Spina ◽  
Alice Pecoraro
2010 ◽  
Vol 134 (3) ◽  
pp. 444-448 ◽  
Author(s):  
Zhengming Gu ◽  
Jianmin Pan ◽  
Matthew J. Bankowski ◽  
Randall T. Hayden

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.


2002 ◽  
Vol 118 (2) ◽  
pp. 550-558 ◽  
Author(s):  
Michael J. Jenner ◽  
Karin E. Summers ◽  
Andrew J. Norton ◽  
John A. Amess ◽  
Rachael S. Arch ◽  
...  

2018 ◽  
Vol 8 (6) ◽  
pp. 554-558 ◽  
Author(s):  
Anne J Blaschke ◽  
Matt McKevitt ◽  
Krow Ampofo ◽  
Tammi Lewis ◽  
Hao Chai ◽  
...  

Abstract Nasopharyngeal (NP) swabs are generally used to detect respiratory syncytial virus (RSV) in infants. However, midturbinate (MT) swabs may provide comparable results. In this study, we enrolled hospitalized infants aged <24 months with RSV and collected NP and MT swabs. The resulting viral loads measured by real-time reverse-transcription quantitative polymerase chain reaction were similar. Most parents preferred MT swabs over NP swabs.


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