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3247X | LIGHT DIAGNOSTICS CMV pp65 Antigenemia IFA Kit, ~125 tests

3247X
5 mL  
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      Key ApplicationsDetection Methods
      ICC Fluorescent
      Description
      Catalogue Number3247X
      Brand Family Chemicon®
      Trade Name
      • LIGHT DIAGNOSTICS
      • Chemicon
      DescriptionLIGHT DIAGNOSTICS CMV pp65 Antigenemia IFA Kit, ~125 tests
      OverviewThe Light Diagnostics Cytomegalovirus (CMV) pp65 Antigenemia Immunofluorescence Assay (IFA) is intended for the qualitative identification of lower matrix protein pp65 of CMV in isolated peripheral blood leukocytes.

      For in vitro Diagnostic use (CE mark)

      Test Principle:

      The Light Diagnostics CMV pp65 Antigenemia Immunofluorescence Assay utilizes an indirect immunofluorescence technique for identifying the lower matrix protein pp65 of human CMV in cytospin preparations of peripheral blood leukocytes. The blend of monoclonal antibodies provided will bind to CMV pp65 antigen present in formalin fixed leukocytes. Unbound monoclonal antibody is removed by washing with phosphate buffered saline (PBS). Fluorescein isothiocyanate (FITC) conjugated antibody will bind to the antigen-antibody complex. Unbound conjugate is removed by washing with PBS. FITC exhibits an apple green fluorescence when excited by ultraviolet light allowing visualization of the complex by fluorescence microscopy. Nuclear fluorescence indicates a positive specimen. Uninfected cells counterstain dull red due to the presence of Evans blue in the FITC conjugated antibody reagent.

      Summary And Explanation:

      CMV is a member of the family Herpesviridae. Synthesis of viral DNA and assembly of capsids occur in the nucleus where infective progeny are released by budding through the nuclear envelope. Also characteristic of herpesviruses, CMV undergoes periods of latency and reactivation.

      CMV is species specific and has been isolated from many animal species (1). The first human CMV was isolated in 1956 from embryonic fibroblasts of adenoidal tissue (2). The terminal morphology of CMV infected permissive cells is that of a large cell with a prominent intranuclear inclusion (Cowdry Type A or "owl eye" cell). Such cells were identified in tissues of fatally infected infants which gave rise to the name "cytomegalic inclusion disease (CID)(3).

      Humans are believed to be the only reservoir of human CMV and postnatal infections are acquired by close contact with individuals shedding virus. Transmission may be through saliva, urine, cervical and vaginal secretions, semen, breast milk, tears, feces, and blood (4,5,6). Infection rates vary with geographic location and socioeconomic conditions.

      CMV infection has been detected in newborn infants and is the most commonly identified cause of congenital infection. Those infants that develop symptoms may exhibit severe disease with jaundice, hepatosplenomegaly, petechiae, and central nervous system abnormalities. The risk of infection is probably the same throughout pregnancy; CID occurs most often in fetuses infected during the first half of gestation. CMV may be transmitted to about 50% of the fetuses after primary maternal infection and about 10% of these will be clinically affected (7,8). Many congenitally infected infants appear normal at birth but subsequently develop neurologic sequelae. The route of transmission of CMV from mother to fetus has not been well elucidated. It is possible that the spread is hematogenous through cord blood or placental tissue and amniotic cells.

      During the past decade an increasing population of immunosuppressed individuals has resulted in a resurgence of CMV as a major pathogen. Induced immunosuppression has occurred more frequently via chemotherapy and transplant regimens. CMV infection is common in patients receiving renal (9), bone marrow (10), heart (11), lung (12), and liver transplants (13). The spread of acquired immunodeficiency syndrome (AIDS) is also related to the CMV resurgence. CMV is the most common opportunistic viral infection in AIDS patients (14,15) and has been implicated as a cofactor in the pathogenesis of the HIV virus (16). CMV infection in AIDS has been implicated in pneumonitis (17), colitis (18,19), retinitis (20), and dementia (21).

      Detection of CMV in blood leukocytes is closely associated with the clinical manifestations of CMV disease and is useful in the diagnosis of CMV infection (22). Rapid diagnosis of CMV disease may prevent delay in treatment using antiviral drugs such as ganciclovir and foscarnet. The Light Diagnostics CMV pp65 Antigenemia Immunofluorescence Assay is a rapid, sensitive method for detection of CMV in isolated leukocytes.
      Materials Required but Not DeliveredEquipment and Reagents:

      · Cytocentrifuge (Shandon Lipshaw, Cytospin 3) cytocentrifuge slides, holders, funnels and filter cards.

      · Laboratory centrifuge

      · Fluorescence microscope with appropriate filter combination for FITC (excitation = 490 nm, emission = 515 nm) with 100x, 200x, and 400x magnification (dry objective)

      · Hemocytometer or Coulter counter

      · No. 1 microscope slide coverslips

      · Sodium hypochlorite solution, 0.05% (1:100 dilution of household bleach)

      · Humid Chamber

      · Incubator (37oC)

      · Coplin staining jars

      · Filter sterilized deionized or distilled water

      · 0.2 M filtered deionized or distilled water
      References
      Product Information
      Components
      • CMV pp65 Monoclonal Antibody - (Catalog No. 5097) One dropper vial containing 5 mL of a monoclonal antibody blend against CMV pp65 antigen, protein stabilizer, 0.05% Tween 20, and 0.1% sodium azide.
      • Anti-Mouse IgG:FITC Conjugate - (Catalog No. 5024) One dropper vial containing 10 mL of FITC labeled anti-mouse IgG, 0.02% Evans blue, protein stabilizer and 0.1% sodium azide.
      • Erythrocyte Lysis Buffer (5X) - (Catalog No. 5122-5X) One bottle containing 200 mL of buffered ammonium chloride. (NH4Cl).
      • Fixation Solution (5X) - (Catalog No. 5113) One bottle containing 220 mL of PBS, formalin, sucrose and 0.5% sodium azide.
      • Permeabilization Solution (5X) - (Catalog No. 5115) One bottle containing 220 mL of PBS, Nonidet P-40, sucrose, protein stabilizer and 0.5% sodium azide.
      • Phosphate Buffered Saline (PBS) - (Catalog No. 5087) Three packets of phosphate buffered saline salts.
      • Mounting Fluid - (Catalog No. 5013) One dropper bottle containing 10 mL of Tris buffered glycerin, a fluorescence enhancer and 0.1% sodium azide.
      Detection methodFluorescent
      Applications
      Key Applications
      • Immunocytochemistry
      Biological Information
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Usage Statement
      • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
      • For in vitro Diagnostic Use
      • CE Mark
      Storage and Shipping Information
      Storage ConditionsWhen stored at 2-8°C, the CMV kit is stable up to the expiration date printed on the kit label. Do not freeze or expose to elevated temperatures. Discard any remaining reagents after the kit expiration date.

      Warnings and Precautions:

      * The sodium azide (NaN3) used as a preservative is toxic if ingested. Sodium azide may react with lead and copper plumbing to form highly explosive metal azides (23, 24). Upon disposal, flush with large volumes of water to prevent build-up in plumbing.

      * Pooling or alteration of any reagent may cause erroneous results.

      * Do not mix or substitute reagents from other manufacturers.

      * Do not allow the slides to dry at any time during the staining procedure.

      * Handle all specimens and materials coming in contact with them as potentially infectious and dispose of with proper precautions. Decontaminate with 0.05% sodium hypochlorite.

      · Formaldehyde solution causes irritation of skin, eyes, nose, and throat. Avoid prolonged or repeated contact. Avoid prolonged breathing of vapor. Use adequate ventilation.

      · Avoid contact with Evans blue (present in the Anti-mouse IgG:FITC conjugate) as it is a potential carcinogen. If skin contact occurs, flush with large volumes of water.

      · Avoid contact with the Mounting Fluid which contains a fluorescence enhancer that may be destructive to tissue of the mucous membranes. If contact occurs, flush with large volumes of water.

      · Incubation times or temperatures other than those specified may give erroneous results. Any such change must be validated by the user.

      · Performance of the fluorescence microscope is of critical importance in achieving satisfactory test results. Microscope objectives, bulb intensity and wattage and filters may affect results.

      * Do not mouth pipette reagents.
      Packaging Information
      Material Size5 mL
      Transport Information
      Supplemental Information
      Specifications