Abstract:
:One of the most interesting aspects of real-time PCR based on the detection of fluorophoric labeled oligonucleotides is the possibility of being able to detect conveniently multiple targets in the same PCR reaction. Recently, Roche Diagnostics launched a real-time PCR platform, the LightCycler480 (LC480), which should be well suited for multiplex real-time PCR analysis. In this paper the performance of the LC480 and accompanying software for the detection of five different targets was analyzed. Target DNAs mixed at equimolar concentrations were detected reproducibly and quantitatively. In addition, mixing different concentrations of the five targets demonstrated that the LC480 is capable of providing quantitative results for a mixture of DNA sequences without losing sensitivity. When applied to the practice of molecular diagnosis of four respiratory viral infections the multiplex assay showed almost complete concordance with corresponding single-target PCRs. The application of multiplex PCR for the detection of multiple pathogens within the same sample will provide a major contribution to the efficiency, logistics and cost-effectiveness of molecular diagnostics.
journal_name
J Virol Methodsjournal_title
Journal of virological methodsauthors
Molenkamp R,van der Ham A,Schinkel J,Beld Mdoi
10.1016/j.jviromet.2006.12.007subject
Has Abstractpub_date
2007-05-01 00:00:00pages
205-11issue
2eissn
0166-0934issn
1879-0984pii
S0166-0934(06)00452-6journal_volume
141pub_type
杂志文章abstract::Hepatitis C virus (HCV) entry into target cells is thought to be a multistep process involving several cellular factors. However, their precise role during virus entry is unclear. Investigation of the mechanisms of HCV entry, such as the order of intervention by the cellular receptors, requires synchronizing infection...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2008.11.015
更新日期:2009-04-01 00:00:00
abstract::The Staphylococcus protein A coagglutination test (Sp A COAT) was developed for the diagnosis of rabbit haemorrhagic disease (RHD). Liver, spleen, lungs and kidneys from 176 rabbits dead from viral haemorrhagic disease as well as the same organs from 64 healthy animals were examined by Sp A COAT and haemaglutination t...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/0166-0934(95)01914-6
更新日期:1996-03-01 00:00:00
abstract::Rubella virosomes were prepared from performed liposomes and detergent solubilized viral hemagglutinin. The liposomes were made from lecithin/dicetyl phosphate (3.5 : 1) films resuspended in NTE buffer and sonicated. Viral hemagglutinin was prepared from purified virus after solubilization with Triton X-100 and centri...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/0166-0934(81)90069-0
更新日期:1981-11-01 00:00:00
abstract::CD4 T-cell count is a priority for staging HIV disease and guiding clinical management as part of HIV care. Conventional CD4 T-cell enumeration methods based on flow cytometry are expensive, require well-trained personnel, and are challenging to use in rural, resource-scarce areas. A simple CD4 T-cell count test that ...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2019.05.012
更新日期:2019-09-01 00:00:00
abstract::Recombinant plasmids carrying 199 base pairs (bp) inserts from the non coding region (nucleotides (nt) 6-204) of the TT virus (TTV) genome were used to standardize an heteroduplex mobility assay able to detect mixed infections of a single individual with several TTV isolates. In this simplified heteroduplex mobility a...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/s0166-0934(01)00425-6
更新日期:2002-03-01 00:00:00
abstract::The Roche Amplicor PCR kit was used to detect HIV-1 DNA in UK patients of known serostatus. Four false-negative and/or equivocal results were obtained from patients who were known to be anti HIV seropositive (Tosswill et al., 1994). Cells from the blood of these patients were shown to contain HIV DNA after extraction,...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/0166-0934(94)00139-8
更新日期:1995-03-01 00:00:00
abstract::A modified sandwich enzyme-linked immunosorbent assay using biotin-streptavidin system (BS-ELISA) was developed to determine levels of tumor necrosis factor-alpha (TNF-alpha) in serum samples of children infected with dengue virus (n=99) and healthy controls (n=41). The minimum detectable concentration of TNF-alpha by...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/s0166-0934(00)00215-9
更新日期:2000-10-01 00:00:00
abstract::Concurrent infection of pigs with two or more pathogens is common in pigs under intensive rearing conditions. Porcine circovirus type 2 (PCV2), porcine parvovirus (PPV), porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV), Japanese encephalitis virus (JEV) and pseudorabies v...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2014.06.027
更新日期:2014-11-01 00:00:00
abstract::Feline calicivirus (FCV) has been used by researchers as a surrogate for Norwalk virus (NV), since they share a similar genomic organization, physicochemical characteristics, and are grouped in the same family, Caliciviridae. Unlike NV, however, FCV can grow in established cell lines and produce a syncytial form of cy...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/s0166-0934(02)00214-8
更新日期:2003-02-01 00:00:00
abstract::The open reading frame coding for the A-type inclusion body protein (ATI) of monkeypox virus (MPV) was identified and sequenced for two strains. Nucleotide sequence comparison revealed 72-95.3% homology with the reported open reading frame sequences of the ATIs of other orthopoxvirus species, such as variola, vaccinia...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/s0166-0934(98)00099-8
更新日期:1998-10-01 00:00:00
abstract::The geographical distribution of human immunodeficiency virus type 1 (HIV-1) subtypes show, with the exception of some African Countries, a specific pattern. However, the significant phenomenon of migration to Western Countries, coupled to inter-ethnic blending, may result in a constant introduction and spread of nove...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2004.11.014
更新日期:2005-03-01 00:00:00
abstract::Serologic diagnosis of human respiratory syncytial virus (hRSV) and human metapneumovirus (hMPV) infections has been shown to complement virus detection methods in epidemiologic studies. Enzyme immunoassays (EIAs) using cultured virus lysate antigens are often used to diagnose infection by demonstration of a ≥4-fold r...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2014.05.008
更新日期:2014-09-01 00:00:00
abstract::To establish a renewable source of parvovirus B19 antigens for diagnostic tests, gene sequences for the viral capsid proteins, VP1 and VP2, were cloned into baculovirus expression vectors and the recombinant viruses used to infect Sf9 insect cells. Cell lysates examined by immunoblotting demonstrated reactive proteins...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/0166-0934(95)00046-w
更新日期:1995-09-01 00:00:00
abstract::Tomato necrotic stunt virus (ToNStV) is an emerging potyvirus that causes severe stunting to infected tomato plants. A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for sensitive detection of ToNStV. The sensitivity of RT-LAMP was comparable to that of conventional RT-PCR, ...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2014.01.017
更新日期:2014-05-01 00:00:00
abstract::This study assesses the ability of quantitative real-time PCR to measure the effects of virus DNA polymerase inhibitors on EBV DNA and late mRNAs syntheses in EBV-producing cell lines. In-house real-time quantitative PCRs were used to measure EBV DNA (thymidine kinase) and mRNAs (BLLF1 gene/gp350/220, BVRF2 gene/prote...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2007.02.005
更新日期:2007-07-01 00:00:00
abstract::Lymphocystis disease virus (LCDV) is the causative agent of lymphocystis disease affecting marine and freshwater fish worldwide. Here an antibody microarray was developed and employed to detect LCDV in fish. Rabbit anti-LCDV serum was arrayed on agarose gel-modified slides as capture antibody, and Cy3-conjugated anti-...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2013.02.015
更新日期:2013-05-01 00:00:00
abstract::Rapid amplification of cDNA ends (RACE) is a powerful PCR-based technique for determination of RNA terminal sequences. However, most of the RACE methods reported in the literature are developed specifically for the mapping of eukaryotic transcripts with 3' poly-A tail and 5' cap structure. In this study, an improved R...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2005.06.022
更新日期:2005-12-01 00:00:00
abstract::A microtiter cell-culture assay is described for measuring neutralizing antibody activity directed against the human immunodeficiency virus (HIV). The assay relies upon inhibition of HIV-mediated cell killing of infected MT-4 lymphoid cells. The assay exhibits comparable sensitivity to two other methods used for such ...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/0166-0934(88)90123-1
更新日期:1988-07-01 00:00:00
abstract::Hepatitis C virus (HCV) infection is a major worldwide health problem, causing chronic hepatitis, liver cirrhosis and primary liver cancer (Hepatocellular carcinoma). HCV encodes a precursor polyprotein that is enzymatically cleaved to release the individual viral proteins. The viral non-structural proteins are cleave...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/s0166-0934(02)00255-0
更新日期:2003-02-01 00:00:00
abstract::Human herpesviruses 6 (HHV-6), 7 (HHV-7) and 8 (HHV-8) are lymphotropic herpesviruses that may cause opportunistic diseases in immunosuppressed patients such as transplant or AIDS patients. The new commercial CMV HHV-6, 7, 8 R-gene kit (Argene, Varilhes, France) for the simultaneous quantitation of HHV-6 and qualitati...
journal_title:Journal of virological methods
pub_type: 杂志文章,多中心研究
doi:10.1016/j.jviromet.2008.01.026
更新日期:2008-05-01 00:00:00
abstract::A simple, and sensitive method for the detection of human papillomavirus (HPV) 16 E6/E7 transcripts in RNA purified from formalin acetic alcohol (FAA)-fixed, paraffin embedded cervical cancer (CaCx) tissue is described. The entire procedure, including polymerase chain reaction (PCR) amplification of cDNA obtained from...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/0166-0934(91)90072-8
更新日期:1991-12-01 00:00:00
abstract::HIV drug resistance (DR) testing using Sanger sequencing (SS) is limited by the inability of the method to identify low abundance drug resistance variants. The application of tagged pooled pyrosequencing (TPP) for HIV DR surveillance is described and the results compared with SS. HIV(+) serum specimens were genotyped ...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2012.10.018
更新日期:2013-02-01 00:00:00
abstract::Cellular nucleic acids can interfere with the molecular cloning of retroviruses, a problem that is particularly serious with viruses propagated in lymphoblastoid cells that release large amounts of microvesicles and other cellular components. The approach taken to circumvent such problems involved first suspending vir...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2006.07.003
更新日期:2006-11-01 00:00:00
abstract::In 2013, a unique seventh yellow head virus genotype (YHV7) was detected in Black Tiger shrimp (Penaeus monodon) broodstock that suffered high mortality following their capture from Joseph Bonaparte Gulf (JBG) in northern Australia. To assist with its diagnosis and assessment of its distribution, prevalence and pathog...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2019.113689
更新日期:2019-11-01 00:00:00
abstract::Tomato ringspot virus (ToRSV), Tobacco ringspot virus (TRSV) and Tobacco rattle virus (TRV) are transmitted to healthy plants by viruliferous nematodes in the soil. A method was developed for extraction of genomic viral RNA from virus particles carried within nematodes and a sensitive nested RT-PCR detection assay. Th...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2008.10.002
更新日期:2009-01-01 00:00:00
abstract::One of the most important steps when preparing a live attenuated vaccine is the assessment of the level of attenuation in target animals. It is costly and time consuming as it requires, on each occasion, a large number of susceptible animals and contained accommodation. This study assessed the consistency of the bovin...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2008.07.007
更新日期:2008-11-01 00:00:00
abstract::The role of environmental reservoirs in avian influenza virus (AIV) transmission has been investigated during AIV-associated outbreaks. To date, no method has been defined for detection of AIV from mud samples. A procedure using elution and polyethylene glycol (PEG) concentration steps was designed to detect AIV by RT...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2011.10.013
更新日期:2012-01-01 00:00:00
abstract::Peste des petits ruminants (PPR) is a viral disease of small ruminants that is caused by the PPR virus (PPRV) and is a significant burden on subsistence farmers across the developing world. Loop-mediated isothermal amplification (LAMP) provides cost-effective, rapid, specific and sensitive detection of nucleic acid an...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2019.113730
更新日期:2019-12-01 00:00:00
abstract::Modification of an in situ polymerase chain reaction (ISPCR) technique is described for the detection of B19 parvovirus infection. Specific amplification of B19 DNA inside fixed cells was followed by hybridisation with a digoxigenin-labelled probe and then visualised by immunochemical reaction. The assay had higher se...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/0166-0934(94)90164-3
更新日期:1994-12-01 00:00:00
abstract::Participation in external quality assessment programs is critical to ensure quality clinical laboratory testing. Commercially available proficiency test panels for HIV-1 virus load testing that are used commonly in external quality assessment programs remain a financial obstacle to resource-limited countries. Maintain...
journal_title:Journal of virological methods
pub_type: 杂志文章
doi:10.1016/j.jviromet.2012.11.036
更新日期:2013-03-01 00:00:00