1,721,347 research outputs found
Enumeration and Characterization of Human Memory T Cells by Enzyme-Linked Immunospot Assays
Full text linkThe enzyme-linked immunospot (ELISPOT) assay has advanced into a useful and widely applicable tool for the evaluation of T-cell responses in both humans and animal models of diseases and/or vaccine candidates. Using synthetic peptides (either individually or as overlapping peptide mixtures) or whole antigens, total lymphocyte or isolated T-cell subset responses can be assessed either after short-term stimulation (standard ELISPOT) or after their expansion during a 10-day culture (cultured ELISPOT). Both assays detect different antigen-specific immune responses allowing the analysis of effector memory T cells and central memory T cells. This paper describes the principle of ELISPOT assays and discusses their application in the evaluation of immune correlates of clinical interest with a focus on the vaccine field
Comparison of three different methods for the evaluation of IL28 and ITPA polymorphisms in patients infected with HCV.
No full textA single nucleotide polymorphism (SNP) upstream of the 1128 gene (rs12979860) has been reported to predict sustained virological response to peginterferon-ribavirin therapy in chronic HCV patients. In addition, two functionally deficient variants (rs1127354 and rs7270101) of inosine triphosphatase (ITPA) were shown to protect against ribavirin (RBV) - induced hemolytic anemia during early stages of treatment. In this study, three methods for detecting IL28 and ITPA mutations were compared to evaluate accuracy, sensitivity costs and turn-around time. IL28 and ITPA variants were detected using genomic DNA from peripheral blood mononuclear cells (PBMCs) of 61 patients with chronic HCV infection by denaturing high-performance liquid chromatography (DHPLC), direct DNA sequencing analysis and Taq Man Real-Time SNP analysis. Complete concordance in the IL28 polymorphism analysis was observed among the three methods. As for ITPA polymorphisms, 60/61 (98.4%) samples were consistent among the three methods, while results for 1/61 (1.64%) samples were concordant by DHPLC and sequencing, and discordant by real-time SNP. All three methods are suitable for routine testing. On the other hand, screening by real-time SNP detection was less expensive and more rapid
Letter to the editor. Virological analysis of fatal influenza cases in the United Kingdom during the early wave of influenza in winter 2010/11
No full textEvaluation of Xpert® Norovirus Assay performance in comparison with real-time RT-PCR in hospitalized adult patients with acute gastroenteritis
No full textXpert® Norovirus Assay (Cepheid, Sunnyvale, CA) was compared with a laboratory-developed real-time RT-PCR assay for the detection of Norovirus GI and GII in hospitalized patients with acute gastroenteritis. The two assays showed a high level of concordance but Xpert® Norovirus Assay allowed faster detection of Norovirus and a simpler sample handling
A new Real-Time RT-PCR Assay for Detection of Human Enterovirus 68 (EV-D68) in Respiratory Samples.
No full textA global reemergence of human enterovirus 68 (EV-D68) associated with severe respiratory illness was occurred during the 2014. We developed and validated an EV-D68-specific real-time RT-PCR for the detection of EV-D68 in respiratory samples. The rapid diagnosis of EV-D68 may contribute to better management of EV-D68 infections
Zika virus isolation from semen
No full textZika virus (ZIKV) can be sexually transmitted and replicative particles were first detected in a semen sample from a patient during the 2013-14 French Polynesia outbreak. Here we describe the virus isolation from semen of a patient returning to Italy from Brazil
Xenotropic and polytropic murine leukemia virus-related sequences are not detected in the majority of patients with chronic fatigue syndrome.
No full textXMRV and polytropic MLV-related virus have been controversially associated with chronic fatigue syndrome (CFS). Subsequent reports failed to detect XMRV and MLV-related virus in CFS patients, and the previous results have been interpreted as a massive laboratory contamination by mouse DNA sequences. Among 12 sequental CFS patients, two were positive for XMRV/MLV sequences. In contrast, 40 selected control subjects were negative CSF patients and controls were negative for mitochondrial mous-specific DNA sequences. These findings do not confirm the high frequency of MLV-related viruses infeciton in CFS patients, but also contrast the widespread laboratory contamination previously suggested
Cellular DNA quantification in respiratory samples for the normalization of viral load: a real need?
No full textBackground: Respiratory tract infections have an enormous social economic impact, with high incidence of hospitalization and high costs. Adequate specimen collection is the first crucial step for the correct diagnosis of viral respiratory infections.
Objectives: The present retrospective study aimed: i) to verify the cell yield obtained from sampling the nasal respiratory tract using mid-turbinate flocked swabs; ii) to evaluate the normalization of viral load, based on cell number; and iii) to compare the kinetics of viral infection obtained with normalized vs non-normalized viral load.
Study design: The number of cells were quantified by real-time PCR in residual extract of nasal swabs tested for respiratory viruses detection and stored at - 80 degrees C in a universal transport medium (UTM (TM)).
Results: A total of 513 virus-positive and 226 virus-negative samples were analyzed. Overall, a median of 4.42 log(10) beta 2-microgolubin DNA copy number/ml of UTM (TM) (range 1.17-7.26) was detected. A significantly higher number of cells was observed in virus-positive as compared to virus-negative samples (4.75 vs 3.76; p < 0.001). Viral loads expressed as log(10) RNA copies/ml of UTM (TM) and log log(10) RNA copies/median number of cells were compared in virus-positive samples and a strict correlation (r = 0.89, p < 0.001) and agreement (R2 = 0.82) were observed. In addition, infection kinetics were compared using the two methods with a follow-up series of eight episodes of viral infection and the mean difference was -0.57 log(10) (range - 1.99 to 0.40).
Conclusions: The normalization of viral load using cellular load compliments the validation of real-time PCR results in the diagnosis of respiratory viruses but is not strictly needed
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