Laboratory Diagnosis Of Viral Infections
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Laboratory Diagnosis Of Viral Infections
In the diagnostic laboratory, virus infections can be confirmed by a myriad of methods. Diagnostic virology has changed rapidly due to the advent of molecular techniques and increased clinical sensitivity of serological assays. Sampling A wide variety of samples can be used for virological testing. The type of sample sent to the laboratory often depends on the type of viral infection being diagnosed and the test required. Proper sampling technique is essential to avoid potential pre-analytical errors. For example, different types of samples must be collected in appropriate tubes to maintain the integrity of the sample and stored at appropriate temperatures (usually 4 °C) to preserve the virus and prevent bacterial or fungal growth. Sometimes multiple sites may also be sampled. Types of samples include the following: * Nasopharyngeal swab * Blood * Skin * Sputum, gargles and bronchial washings * Urine * Semen * Faeces * Cerebrospinal fluid * Tissues (biopsies or post-mo ...
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Virus
A virus is a submicroscopic infectious agent that replicates only inside the living cells of an organism. Viruses infect all life forms, from animals and plants to microorganisms, including bacteria and archaea. Since Dmitri Ivanovsky's 1892 article describing a non-bacterial pathogen infecting tobacco plants and the discovery of the tobacco mosaic virus by Martinus Beijerinck in 1898,Dimmock p. 4 more than 9,000 virus species have been described in detail of the millions of types of viruses in the environment. Viruses are found in almost every ecosystem on Earth and are the most numerous type of biological entity. The study of viruses is known as virology, a subspeciality of microbiology. When infected, a host cell is often forced to rapidly produce thousands of copies of the original virus. When not inside an infected cell or in the process of infecting a cell, viruses exist in the form of independent particles, or ''virions'', consisting of (i) the genetic material, i. ...
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Hep G2
Hep G2 (or HepG2) is a human liver cancer cell line. Hep G2 is an immortal cell line which was derived in 1975 from the liver tissue of a 15-year-old Caucasian male from Argentina with a well-differentiated hepatocellular carcinoma. These cells are epithelial in morphology, have a modal chromosome number of 55, and are not tumorigenic in nude mice. The cells secrete a variety of major plasma proteins, e.g., albumin, and the acute-phase proteins fibrinogen, alpha 2-macroglobulin, alpha 1-antitrypsin, transferrin and plasminogen. They have been grown successfully in large-scale cultivation systems. Hepatitis B virus surface antigens have not been detected. Hep G2 will respond to stimulation with human growth hormone. Hep G2 cells are a suitable ''in vitro'' model system for the study of polarized human hepatocytes. Another well-characterized polarized hepatocyte cell line is the rat hepatoma-derived hybrid cell line WIF-B. With the proper culture conditions, He ...
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Immunity (medical)
In biology, immunity is the capability of multicellular organisms to resist harmful microorganisms. Immunity involves both specific and nonspecific components. The nonspecific components act as barriers or eliminators of a wide range of pathogens irrespective of their antigenic make-up. Other components of the immune system adapt themselves to each new disease encountered and can generate pathogen-specific immunity. Immunity is a complex biological system that can recognize and tolerate whatever belongs to the self, and to recognize and reject what is foreign (non-self). Innate and adaptive Innate Immunity First off, the immune system is a system in the body of animals that enables animals to avoid or limit many infections caused by pathogens. Pathogens are disease causing agents, causing a wide range of illnesses. As for Diseases it's when structure of an organism is negatively affected other than external injury. Both diseases and pathogens affect the immune system causing ill ...
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Immunoglobulin G
Immunoglobulin G (Ig G) is a type of antibody. Representing approximately 75% of serum antibodies in humans, IgG is the most common type of antibody found in blood circulation. IgG molecules are created and released by plasma B cells. Each IgG antibody has two paratopes. Function Antibodies are major components of humoral immunity. IgG is the main type of antibody found in blood and extracellular fluid, allowing it to control infection of body tissues. By binding many kinds of pathogens such as viruses, bacteria, and fungi, IgG protects the body from infection. It does this through several mechanisms: * IgG-mediated binding of pathogens causes their immobilization and binding together via agglutination; IgG coating of pathogen surfaces (known as opsonization) allows their recognition and ingestion by phagocytic immune cells leading to the elimination of the pathogen itself; * IgG activates all the classical pathway of the complement system, a cascade of immune protein pr ...
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Humoral Immunity
Humoral immunity is the aspect of immunity that is mediated by macromolecules - including secreted antibodies, complement proteins, and certain antimicrobial peptides - located in extracellular fluids. Humoral immunity is named so because it involves substances found in the humors, or body fluids. It contrasts with cell-mediated immunity. Humoral immunity is also referred to as antibody-mediated immunity. The study of the molecular and cellular components that form the immune system, including their function and interaction, is the central science of immunology. The immune system is divided into a more primitive innate immune system and an acquired or adaptive immune system of vertebrates, each of which contain both humoral and cellular immune elements. Humoral immunity refers to antibody production and the coinciding processes that accompany it, including: Th2 activation and cytokine production, germinal center formation and isotype switching, and affinity maturation an ...
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Immunoperoxidase
Immunoperoxidase is a type of immunostain used in molecular biology, medical research, and clinical diagnostics. In particular, immunoperoxidase reactions refer to a sub-class of immunohistochemical or immunocytochemical procedures in which the antibodies are visualized via a peroxidase-catalyzed reaction. Immunohistochemistry and immunocytochemistry are methods used to determine in which cells or parts of cells a particular protein or other macromolecule are located. These stains use ''antibodies'' to bind to specific ''antigens'', usually of protein or glycoprotein origin. Since antibodies are normally invisible, special strategies must be employed to detect these bound antibodies. In an immunoperoxidase procedure, an enzyme known as a peroxidase is used to catalyze a chemical reaction to produce a coloured product. Simply, a very thin slice of tissue is fixed onto glass, incubated with antibody or a series of antibodies, the last of which is chemically linked to peroxidase. Aft ...
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Immunofluorescence
Immunofluorescence is a technique used for light microscopy with a fluorescence microscope and is used primarily on microbiological samples. This technique uses the specificity of antibodies to their antigen to target fluorescent dyes to specific biomolecule targets within a cell, and therefore allows visualization of the distribution of the target molecule through the sample. The specific region an antibody recognizes on an antigen is called an epitope. There have been efforts in epitope mapping since many antibodies can bind the same epitope and levels of binding between antibodies that recognize the same epitope can vary. Additionally, the binding of the fluorophore to the antibody itself cannot interfere with the immunological specificity of the antibody or the binding capacity of its antigen. Immunofluorescence is a widely used example of immunostaining (using antibodies to stain proteins) and is a specific example of immunohistochemistry (the use of the antibody-antigen rel ...
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Antiviral Therapy
Antiviral drugs are a class of medication used for treating viral infections. Most antivirals target specific viruses, while a broad-spectrum antiviral is effective against a wide range of viruses. Unlike most antibiotics, antiviral drugs do not destroy their target pathogen; instead they inhibit its development. Antiviral drugs are one class of antimicrobials, a larger group which also includes antibiotic (also termed antibacterial), antifungal and antiparasitic drugs, or antiviral drugs based on monoclonal antibodies. Most antivirals are considered relatively harmless to the host, and therefore can be used to treat infections. They should be distinguished from viricides, which are not medication but deactivate or destroy virus particles, either inside or outside the body. Natural viricides are produced by some plants such as eucalyptus and Australian tea trees. Medical uses Most of the antiviral drugs now available are designed to help deal with HIV, herpes viruses, the he ...
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Real Time PCR
A real-time polymerase chain reaction (real-time PCR, or qPCR) is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR). It monitors the amplification of a targeted DNA molecule during the PCR (i.e., in real time), not at its end, as in conventional PCR. Real-time PCR can be used quantitatively (quantitative real-time PCR) and semi-quantitatively (i.e., above/below a certain amount of DNA molecules) (semi-quantitative real-time PCR). Two common methods for the detection of PCR products in real-time PCR are (1) non-specific fluorescent dyes that intercalate with any double-stranded DNA and (2) sequence-specific DNA probes consisting of oligonucleotides that are labelled with a fluorescent reporter, which permits detection only after hybridization of the probe with its complementary sequence. The Minimum Information for Publication of Quantitative Real-Time PCR Experiments ( MIQE) guidelines propose that the abbreviation ''qPCR'' be used for q ...
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Reverse Transcriptase PCR
Reverse transcription polymerase chain reaction (RT-PCR) is a laboratory technique combining reverse transcription of RNA into DNA (in this context called complementary DNA or cDNA) and amplification of specific DNA targets using polymerase chain reaction (PCR). It is primarily used to measure the amount of a specific RNA. This is achieved by monitoring the amplification reaction using fluorescence, a technique called real-time PCR or quantitative PCR (qPCR). Combined RT-PCR and qPCR are routinely used for analysis of gene expression and quantification of viral RNA in research and clinical settings. The close association between RT-PCR and qPCR has led to metonymic use of the term qPCR to mean RT-PCR. Such use may be confusing, as RT-PCR can be used without qPCR, for example to enable molecular cloning, sequencing or simple detection of RNA. Conversely, qPCR may be used without RT-PCR, for example to quantify the copy number of a specific piece of DNA. Nomenclature The combined ...
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Polymerase Chain Reaction
The polymerase chain reaction (PCR) is a method widely used to rapidly make millions to billions of copies (complete or partial) of a specific DNA sample, allowing scientists to take a very small sample of DNA and amplify it (or a part of it) to a large enough amount to study in detail. PCR was invented in 1983 by the American biochemist Kary Mullis at Cetus Corporation; Mullis and biochemist Michael Smith (chemist), Michael Smith, who had developed other essential ways of manipulating DNA, were jointly awarded the Nobel Prize in Chemistry in 1993. PCR is fundamental to many of the procedures used in genetic testing and research, including analysis of Ancient DNA, ancient samples of DNA and identification of infectious agents. Using PCR, copies of very small amounts of DNA sequences are exponentially amplified in a series of cycles of temperature changes. PCR is now a common and often indispensable technique used in medical laboratory research for a broad variety of applications ...
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