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Hep AD38 Assay: A High-Throughput, Cell-Based Screen for the Evaluation of Compounds Against Hepatitis B Virus

Approximately 5% of the world’s population has been infected with hepatitis B virus (HBV). Ten percent of these adults will become chronic carriers, as will 95% of the infants infected perinatally. Those that do become chronically infected with HBV are at increased risk of developing liver dysf ...

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Hepatitis B Virus Cell Culture Assays for Antiviral Activity

Hepatitis B Virus (HBV) is a causative agent of both acute and chronic hepatitis, a major etiologic factor of primary hepatocellular carcinoma, and a serious global health problem, with over 240 million estimated chronically infected individuals. Although there are several promising ...

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A Rapid Microtiter Assay for Duck Hepatitis Virus Reverse Transcriptase

The duck hepatitis B virus (DHBV) genome contains a pol gene that codes for the viral polymerase protein. This enzyme, which is essential for the replication of the virus, has multiple activities including an RNA directed DNA polymerase or reverse transcriptase (RT) activity, an RNase H activit ...

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Assays for Measuring Hepadnaviral Supercoiled DNA

Amajor reason for treatment failure during antiviral therapy of chronic hepatitis B infection is thought to be the persistence of the key replicative intermediate, the viral covalently closed circular (CCC) or supercoiled DNA (1,2). Investigators studying the structure and functi ...

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Assays for Hepatitis B Virus DNA-and RNA-Dependent DNA Polymerase Activities

Genomes of the hepatitis B viruses (HBVs) consist of approx 3.2 kb of partly double-stranded DNA containing three or four overlapping open reading frames, the largest of which encodes the viral polymerase (Pol) protein. After entry into the cell and uncoating, the viral genome is transported to t ...

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Determination of Viral Infectivity: HSV, VZV, and CMV

Compared to the wide range of antibiotics that are available, the number of antiviral drugs is limited. However, herpesviruses have always been a major target for antiviral drug design, and there are a wide range of drugs at various stages of development. All of the assays described in this chapter pr ...

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Development of a Hepatitis C Virus RNA Helicase High Throughput Assay

In this chapter, the development of a 96-well plate increasing signal helicase assay will be described. The authors have used this assay to detect inhibitors of hepatitis C virus (HCV) NS3/4A RNA helicase.

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Antiviral Screening Assays for Human Herpesviruses 6 and 7

Human herpesviruses 6 and 7 are ubiquitous herpesviruses that normally infect their hosts early in life. There are two variant groups of human herpesvirus 6 (HHV-6): variants A (HHV-6A) and B (HHV-6B). Variant A has not been unambiguously associated with a specific disease but may contribute to di ...

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Strategies for Evaluation of Antiviral Agents Against Epstein-Barr Virus in Culture

Evaluation of antiviral agents against Epstein-Barr virus (EBV) has been hampered by the lack of a permissive cell system for the replication of this virus. The extent of EBV replication detected in producer cell lines (P3HR-1 and B95-8) is limited: only a small fraction (

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Herpesvirus Protease Assays

Herpesviruses encode a serine protease that is essential for the maturation of viral capsids (1,2). The protease is expressed as part of a polyprotein. The catalytic domain is contained within the N-terminal third of the protein, and the remainder comprises a structural “scaffold” protein. T ...

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Assessment of Activity of Topical Virucidal Agents

There is currently considerable interest in the possibility of developing a potent, nontoxic anti-HIV agent that could be used intravaginally to reduce the risks of transmission of HIV. Worldwide up to 80% of HIV infections have been acquired heterosexually. Projections suggest that by t ...

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Evaluation of Compounds That Prevent Reactivation of HIV-1 in OM-10.1 Cells

It is critically important that new therapeutic compounds and targets for therapeutic intervention be identified in the battle against the human immunodeficieny virus type 1(HIV-1). Many of the currently existing therapeutic approaches target virus-specific factors involv ...

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Testing Compounds for Antiviral Activity in Cell Cultures Infected with HIV

This chapter describes the procedures that can be used to determine compounds that have antiviral activity against HIV. These include: maintenance of lymphoblastoid cell lines, preparation of peripheral blood mononuclear cells (PMMCs), and determination of the infectivity of the ...

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Assays That Measure Selective Killing of Virus-Infected Cells

When evaluating potential antiviral compounds, determining the effect of the drug on virus replication is usually the prime concern. However, since virus replication is dependent on the host cell, in circumstances where virus infection is not rapidly lytic to cells, the effect of these com ...

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Evaluating Clinical Isolates for Their Phenotypic and Genotypic Resistance Against Anti-HIV Drugs

The high replication rate of HIV, together with the low fidelity of its reverse transcriptase, provides the virus with an unprecedented genomic flexibility. This allows a fast adaptation to selective pressure, including antiviral drugs, resulting in the development of drug-resista ...

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Line Probe Assay for Detecting Mutations in HIV-1 Reverse Transcriptase

The human immunodeficiency virus type 1 (HIV-1) belongs to the family of positive-stranded, enveloped RNA viruses with a DNA intermediate step (retroviruses). Because of the lack of fidelity of the reverse transcriptase (RT), the replication is error-prone, and the infection is characte ...

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RT-PCR for Amplification of Specific Fragments of HIV-1 Genome

HIV, the etiologic agent of AIDS, is a retrovirus of the family Lentiviridae, first isolated in 1983 by the group of Luc Montagnier at the Pasteur Institute of Paris (1). In the following years, much effort has been, and still is, focused on the search for antiviral drugs that would help to control the course of the ...

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Evaluation of Compounds Against Recombinant HIV Reverse Transcriptase

Reverse transcriptase (RT) has attracted particular attention as a target enzyme for AIDS chemotherapy, because the enzyme catalyzes a crucial step in the HIV replicative cycle. Effective inhibition of this enzyme prevents the formation of proviral DNA. RT is endowed with three indepen ...

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Ultrasensitive Detection of Reverse Transcriptase Activity by the Amp-RT Assay: Applications to the Measurement of Virus Loads and Phenotypic Drug Res

Retroviruses are widely prevalent among vertebrates and are the causative agents of a variety of diseases in humans and animals including immunodeficiences, leukemias, and lymphomas (1). The retrovirus family is characterized by the presence of virion-associated reverse trans ...

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Reverse Transcriptase Assay Based on Product Enhancement for Assessing the Drug Susceptibility of Retroviruses

All replication-competent retroviruses possess a characteristic enzyme, reverse transcriptase (RT), which is present at 20–70 mol/virus particle (1–3). The enzyme is cleaved, and thereby activated, from an inactive precursor by the action of another retroviral enzyme, the viral pro ...

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