Abstract：
:We describe an in-situ RT-PCR method for the amplification of rhinovirus (RV) in fixed, paraffin-embedded HeLa cells employed as a model for human respiratory epithelium. HeLa cells were infected in-vitro with inocula of rhinovirus-16 ranging from 10(2) to 10(6) 50% tissue culture infective doses (TCID50), incubated for 18 h then fixed and processed into paraffin blocks. Sections of the cell preparation were subjected to standard RT-PCR, in-situ hybridisation (ISH) or in-situ RT-PCR using specific oligonucleotide primers or probes directed against the 5' non-coding region of RV RNA. RT-PCR was found to be capable of detecting RV16 RNA in one 8 microns-thick section of cells infected with the lowest virus titre. ISH using digoxigenin labelled oligonucleotide probes located RV16 signal in the majority of HeLa cells at the highest virus titre, but in few or no cells with the lowest virus titre. In contrast, in-situ RT-PCR detected RV16 in the majority of cells infected with this amount of RV16. There was a slight loss of morphology and fine localisation associated with the in-situ thermal cycling process. However, the sensitivity of in-situ RT-PCR is comparable to standard RT-PCR and greater than ISH for the detection of RV. In-situ RT-PCR has wide applications for sensitive localization of low copy viral and RNA sequences within cells to investigate the role of viruses in a variety of clinical conditions.
journal_name
J Virol Methodsjournal_title
Journal of virological methodsauthors
Bates PJ,Sanderson G,Holgate ST,Johnston SLdoi
10.1016/s0166-0934(97)00095-5subject
Has Abstractpub_date
1997-09-01 00:00:00pages
153-60issue
2eissn
0166-0934issn
1879-0984pii
S0166-0934(97)00095-5journal_volume
67pub_type
杂志文章abstract：:Monoclonal antibodies (Mabs) against influenza B virus were obtained by immunizing mice with B/Nagasaki/1/87, one of the strains of the B/Victoria group. Immunoprecipitation analysis revealed that individual Mabs precipitated the nucleoprotein (NP), the matrix protein (M) or the hemagglutinin protein (HA). By using th...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(99)00015-4
更新日期：1999-04-01 00:00:00
abstract：:All human herpesviruses cause chronic infections in which latent virus is periodically reactivated. Persistently active infections are uncommon, however, and occur exclusively in individuals whose immune systems fail to control virus multiplication and spread. This paper summarizes the management of these unusual infe...
journal_title：Journal of virological methods
pub_type： 杂志文章,评审
doi：10.1016/0166-0934(88)90076-6
更新日期：1988-09-01 00:00:00
abstract：:Recently, sequences from a putative member of the Flaviviridae, GB virus C (GBV-C), were isolated from the serum of patients with cryptogenic hepatitis. These sequences were 83-99% identical at the nucleotide level. Because of the divergence between these GBV-C isolates, it is likely that the PCR-based detection assay...
journal_title：Journal of virological methods
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doi：10.1016/0166-0934(95)01956-1
更新日期：1996-01-01 00:00:00
abstract：:A real-time PCR assay based on the TaqMan chemistry was developed for reliable detection and quantitation of the squash leaf curl virus (SLCV) in melon and squash plants. This method was highly specific to SLCV and it was about one thousand times more sensitive than the conventional PCR method. The protocol of the rea...
journal_title：Journal of virological methods
pub_type： 杂志文章
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更新日期：2012-02-01 00:00:00
abstract：:The rapid development of new molecular biology methods has improved infectious disease diagnosis, which is increasingly important to clinical management and public health. A wide variety of new methods which are more specific, sensitive and robust, such as combination of PCR and microarray technology, has gradually re...
journal_title：Journal of virological methods
pub_type： 杂志文章
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abstract：:Commercially marketed kits are now available for rapid viral assay of clinical specimens. This study was conducted to determine the suitability of these kits for use in environmental testing. Eight rotavirus kits and one enteric adenovirus kit were screened for sensitivity using simian rotavirus SA11, human rotavirus ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(93)90098-c
更新日期：1993-12-15 00:00:00
abstract：:Long-term viral archives are valuable sources of research data. Each archive can store hundreds of thousands of diverse sample types. In the current era of whole genome sequencing, archived samples become a rich source of evolutionary and epidemiological data that can span years, and even decades. However, the ability...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2019.113777
更新日期：2020-02-01 00:00:00
abstract：:The polymerase chain reaction (PCR) is one of the most efficient techniques for measuring the viral load of HIV-infected samples. Determination of the specificity of PCR products is usually based on Southern blotting and hybridization of the amplified DNA to radioactive oligonucleotide probes specific for sequences co...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(94)90028-0
更新日期：1994-05-01 00:00:00
abstract：:We show that the immuno-gold technique adapted to electron microscopy is a sensitive method for in situ localization of viral proteins in plant cells. Using antisera raised against cauliflower mosaic virus and the viroplasm major protein (VmP) we obtained a selective labelling of the viral particles and of the viropla...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(84)90053-3
更新日期：1984-12-01 00:00:00
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journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(90)90135-3
更新日期：1990-02-01 00:00:00
abstract：:Avian influenza virus (AIV) is an enveloped virus with segmented RNA that belongs to the Orthomyxoviridae. Recently, avian influenza virus isolates have not only posed a significant threat to the poultry industry but also serious public health concerns. The full-length viral hemagglutinin (HA), neuraminidase (NA) or b...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2011.05.006
更新日期：2011-08-01 00:00:00
abstract：:The laboratory diagnosis of hepatitis B virus (HBV) infection is based mainly on serological assays. Yet the detection and quantitation of viral DNA is necessary when addressing directly the question of infectivity or when monitoring the viral load during therapy. Standard hybridization assays allow for exact quantita...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(99)00154-8
更新日期：2000-03-01 00:00:00
abstract：:Human immunodeficiency virus (HIV) encodes a transcription trans-activator (Tat) with an essential role in the transcriptional elongation of viral RNA based on the viral promoter long terminal repeat (LTR). Tat-mediated transcription is conserved and can be distinguished from host transcription, so it is a therapeutic...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2017.08.005
更新日期：2017-11-01 00:00:00
abstract：:The SPF10 PCR targets a conserved 65bp region of the HPV L1 gene for broad-spectrum amplification. The LiPA assay allows subsequent genotyping of the HPV amplicons. This study aims to develop a SPF10 real-time PCR to achieve simultaneous amplification and detection of the HPV target. That way, LiPA analysis of the HPV...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2012.09.013
更新日期：2013-01-01 00:00:00
abstract：:Herpes simplex virus (HSV) isolates were differentiated by immunoblotting analysis using a mixture of polyclonal antisera directed against HSV type 1 (HSV-1) and HSV type 2 (HSV-2) glycoprotein fractions (gB/gC of HSV-1 and gC/gE/84-kDa protein of HSV-2), since the mixed antisera recognized viral polypeptides with dif...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(87)90122-4
更新日期：1987-11-01 00:00:00
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journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(99)00134-2
更新日期：2000-01-01 00:00:00
abstract：:Human cytomegalovirus (HCMV) is the most frequent cause of congenital infection. Diagnosis of this infection is important because 5-17% of asymptomatic infected babies will develop late sequelae and should be followed closely. Most of these children will remain undetected, since screening of all newborns by viral cult...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2005.02.018
更新日期：2005-09-01 00:00:00
abstract：:The major protective antigens of retroviruses are considered to be their glycosylated envelope proteins. However, the methods commonly employed to enrich and purify virus from culture media such as pelleting and density-gradient centrifugation result in a low recovery of the viral external glycoproteins. This is an ob...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(89)90011-6
更新日期：1989-04-01 00:00:00
abstract：:Mungbean yellow mosaic virus-Vigna (MYMV) sequences cloned as partial dimers within the T-DNA of a binary vector were deleted at a high frequency upon conjugal mobilization from Escherichia coli into Agrobacterium tumefaciens. This deletion involving the genome-length viral DNA did not occur when the binary plasmid wa...
journal_title：Journal of virological methods
pub_type： 杂志文章
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更新日期：2006-10-01 00:00:00
abstract：:The clinical and epidemiologic management of the SARS-CoV-2 pandemic is critically dependent on molecular assays with short turn-around time. We validated the novel Xpert Xpress SARS-CoV-2 assay using a commercial nucleic acid testing (Roche Cobas 6800). We found an excellent concordance over a range of SARS-CoV-2 loa...
journal_title：Journal of virological methods
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pub_type： 杂志文章
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更新日期：2006-06-01 00:00:00
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journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(83)90026-5
更新日期：1983-03-01 00:00:00
abstract：:To map the epitopes of VP2 protein of chicken anemia virus (CAV), VP2 was expressed as a fusion protein in Escherichia coli BL21 (DE3). The Western blot demonstrated that recombinant VP2 protein could be recognized by sera of chickens infected with CAV. Female BALB/c mice were immunized with purified recombinant VP2 p...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2007.03.016
更新日期：2007-08-01 00:00:00
abstract：:A real-time RT-PCR assay was developed utilizing dual-labeled fluorescent probes binding to VP4 sequence that are specific to the classical (Cl), variant (V) and very virulent (vv) strains of infectious bursal disease virus (IBDV). The assay was highly sensitive and could detect as little as 3 x 10(2) to 3 x 10(3) cop...
journal_title：Journal of virological methods
pub_type： 杂志文章
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更新日期：2005-07-01 00:00:00
abstract：:Puumala virus, a hantavirus belonging to the Bunyaviridae family, causes a human disease known as nephropathia epidemica, a mild form of hemorrhagic fever with renal syndrome. The implementation of effective decontamination procedures is critical in hantavirus research to minimize the risk of personnel exposure. This ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2006.11.037
更新日期：2007-04-01 00:00:00
abstract：:A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay has been developed to detect and differentiate between canine parvovirus (CPV) and mink enteritis virus (MEV). Eight CPV and three MEV epidemic strains isolated from 28 pathological samples from dogs and minks suspected of being infe...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.09.014
更新日期：2014-12-15 00:00:00
abstract：:Lily symptomless virus (LSV) occurs frequently in many Lilium species worldwide and often causes developmental abnormalities such as a smaller flower and lower bulb yield. In this study, two moderate and efficient gel filtration chromatography (GFC) methods was compared, these two techniques were, respectively, based ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2006.09.008
更新日期：2007-02-01 00:00:00
abstract：:The aim of this study was the development of gag and pol dual labelled probe real-time PCR and RT PCR assays to quantify the proviral load and the transcripts of the British Visna/maedi virus EV1 strain. Primers and probes were chosen based on the consensus sequences of gag and pol clones representative of EV1 genetic...
journal_title：Journal of virological methods
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更新日期：2010-05-01 00:00:00
abstract：:The interplay between avian reovirus (ARV) replication and apoptosis and proteasome pathway was studied in cultured cells. It is shown that inhibition of the proteasome did not affect viral entry and host cell translation but had influence on ARV replication and ARV-induced apoptosis. Evidence is provided to demonstra...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2008.03.016
更新日期：2008-07-01 00:00:00
abstract：:Enzyme immunoassays based on the use of monoclonal antibodies (MAbs) were developed for the detection of the barley stripe mosaic virus (BSMV) and the beet necrotic yellow vein virus (BNYVV). Assays employing conjugates of MAbs to horseradish peroxidase (HRP) were compared to systems with biotinylated MAbs and strepta...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(95)01962-6
更新日期：1996-02-01 00:00:00
