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3108 | LIGHT DIAGNOSTICS™ Respiratory Viral Screen IFA, included in kit #3105

3108
10 mL  
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      Overview

      Replacement Information

      Key Spec Table

      Key ApplicationsFormatHostDetection Methods
      IFPurifiedMFluorescent
      Description
      Catalogue Number3108
      Brand Family Chemicon®
      Trade Name
      • LIGHT DIAGNOSTICS
      • Chemicon
      DescriptionLIGHT DIAGNOSTICS™ Respiratory Viral Screen IFA, included in kit #3105
      OverviewThe Light Diagnostics Respiratory Viral Screen is intended for in vitro diagnostic use in the qualitative confirmation of adenovirus, influenza A, influenza B, parainfluenza 1, parainfluenza 2, parainfluenza 3, and respiratory syncytial virus (RSV) in inoculated cell cultures.

      Summary and Explanation:

      Respiratory viruses are responsible for a significant proportion of illness in human populations. Some viruses (e.g. influenza) are seasonal while others (e.g. adenovirus) predominantly affect different age groups. In a given population, respiratory viruses may be responsible for a considerable amount of morbidity. The advent of appropriate anti-viral therapy against some respiratory viruses makes rapid screening to identify these organisms imperative, allowing early institution of therapy. The two most widely used anti-viral agents today are amantadine/rimantadine for influenza A and ribavirin for respiratory syncytial virus bronchiolitis. Both agents have varying modes of action although most involve early infection steps such as penetration or uncoating40,62. At physiologically attainable concentrations, amantadine and rimantadine specifically inhibit replication of influenza A12. Ribavirin has broad spectrum activity in vitro against a host of viruses such as respiratory syncytial virus, measles, influenza A and B, and parainfluenza45,47,23.

      The three commonly used modes of identifying viruses are: culture isolation/ confirmation, direct detection, and serology. Of these, culture isolation/confirmation is the standard method for most laboratories. It is to date the most sensitive method available for the detection of respiratory viruses and is sometimes used to confirm direct detection findings. Light Diagnostics Respiratory Viral Screen utilizes monoclonal antibodies for culture confirmation to provide clear, easy to interpret results in only 60 minutes.

      Adenoviruses are responsible for a significant number of clinical respiratory illnesses. Upper respiratory diseases caused by adenovirus include colds, pharyngitis, and tonsillitis and occur mostly in infants and young children. Approximately 10% of childhood pneumonia is probably caused by adenovirus44. Other adenovirus related lower respiratory illnesses include bronchitis and bronchiolitis31. In children under 5, adenovirus is responsible for about 5% of cases of acute respiratory disease (ARD). ARD may be manifested by nasal congestion, coryza, cough and, at times, tonsillitis, fever, and myalgia. The appearance of conjunctivitis with ARD constitutes pharyngoconjunctival fever2. Adenovirus has commonly been associated with pertussis syndrome, but recent studies suggest that the presence of adenovirus in these cases may be a reactivation of latent virus from tonsillar tissue during Bordetella pertussis infections53.

      Ocular illnesses resulting from adenovirus infection include epidemic keratoconjunctivitis (EKC), acute hemorrhagic conjunctivitis, and acute follicular conjunctivitis.

      Adenovirus is related to several gastrointestinal disorders and is probably evident in 7-17% of all childhood gastroenteritis. Types 40 and 41 have been associated with diarrhea and acute gastroenteritis13,22,68.

      Adenovirus has also been linked with intussusception3, acute hemorrhagic cystitis54, and meningoencephalitis.

      Research indicates that the incidence of adenovirus infection in immunocompromised patients is probably no higher than in normal individuals, however, severity and probability of death may be greater 31.

      Human adenoviruses belong to the family Adenoviridae, genus Mastadenovirus. They are non-enveloped, double stranded DNA viruses icosahedral in shape ranging from 70-90 nm in diameter30. They have a protein coat of 240 hexon and 12 penton capsomeres.

      Adenovirus can be cultured and isolated in a variety of cell lines and identified by several methods. Suitable cell lines include Hep-2, HeLa, KB, and HEK. Graham 293 cells may be used for propagation of types 40 and 41. Confirmation of infection is usually achieved by immunofluorescence or enzyme immunoassay (EIA), but it can be done by complement fixation, hemagglutination-inhibition, and neutralization methods34.

      Influenza virus causes highly contagious respiratory disease which typically results in epidemics28. There are three types, A, B, and C, in which specificity is conferred by internal nucleoprotein and matrix protein antigens52.

      Adult infection with influenza virus characteristically results in tracheobronchitis and small airway involvement42,65 with possible development of rhinitis and/or pharyngitis. Infection can, however, display clinical manifestations ranging from no symptoms to fatal pneumonia52. The same spectrum of clinical response can be seen in children with some distinct differences. Fever may be higher in children and accompanied with febrile convulsions24,26,60,70,71. Influenza viruses are responsible for 14% of childhood fevers with respiratory tract symptoms severe enough to warrant a physician's attention24,71. Children experience more gastrointestinal involvement than adults14 and they develop myositis, otitis media, and croup more frequently29,38. Neonatal infection may result in unexplained fever48 and is potentially fatal1,18,48,57. Lower respiratory tract disease in children and adults associated with influenza infection is manifested in three forms of pneumonia: primary viral pneumonia, combined viral-bacterial pneumonia, and influenza infection followed by bacterial pneumonia52. Non-pulmonary clinical responses to influenza infection include viremia, cardiac and CNS involvement, Reyes syndrome, and toxic shock52.

      Influenza types A and B cause essentially the same spectrum of disease. Type A infection, however results in hospitalization approximately four times more often than type B38, and type B more commonly results in myositis and gastrointestinal involvement11,15,36,41.

      Influenza type C rarely results in lower respiratory tract illness but causes sporadic upper respiratory tract disease38,19,50. By adulthood, almost all individuals have antibody to type C55.

      Influenza viruses are members of the family Orthomyxoviridae. They are pleomorphic, enveloped and contain a segmented, single-stranded, negative-sense RNA genome. They range in diameter from 80-120nm39.

      Addition of trypsin makes it possible to culture influenza viruses in a variety of cell lines such as Madin Darby canine kidney (MDCK), A549 lung carcinoma and primary monkey kidney (PMK)20,64 as well as in embryonated hen eggs or the allantoin-on-shell system17. Influenza virus can be detected by immunofluorescence, hemadsorption or hemagglutination techniques using chicken or guinea pig erythrocytes. Isolation/culture confirmation provides an easy, sensitive and relatively rapid technique for identifying influenza infections. Cytopathic effect is evinced in 3-7 days post-inoculation as vacuolation and cell degradation.

      Parainfluenza viruses, combined with respiratory syncytial virus, represent the most significant upper respiratory pathogens in infants and young children4,6,7,37. Four types of parainfluenza virus have been identified in children and adults. Types 1 and 2 are major causes of laryngotracheobronchitis (croup). The severity of illness is greatest in children ages 2-4 years66. Parainfluenza type 3 infection can lead to croup but, most notably, type 3 is second only to respiratory syncytial virus as a cause of infant bronchiolitis and pneumonia8,9,25,58,59. Illness from type 3 infection is most severe in infants less than 1 year old66.

      In older children and adults, illness may be asymptomatic or mimic the common cold67. Severe croup in early childhood or infancy may result in bronchial hyperactivity in older children or adolescents after exercise. However, it remains undetermined whether bronchial hyperactivity was a preexisting condition which played a role in the pathogenesis of croup or whether it develops as a complication of severe croup27,43.

      Parainfluenza type 4 has been associated only with mild upper respiratory illness in adults and children and is difficult to identify in cell cultures66.

      Parainfluenza viruses belong to the genus Paramyxovirus of the family Paramyxoviridae. They are enveloped viruses with a single-strand RNA genome of negative polarity and range in diameter from 150-200nm10.

      Parainfluenza viruses grow well in primary simian or human kidney cell lines and in LLC-MK2, a rhesus kidney heteroploid cell line66. Trypsin is needed in the medium for the recovery of types 1 and 2 but not type 3. Virus infection of tissue culture can be recognized by hemadsorption of guinea pig erythrocytes. Types 2 and 3 can be recognized by syncytium formation.

      By age 2, most children have experienced Respiratory Syncytial Virus (RSV) infection, making it the most important viral cause of childhood lower respiratory tract illness63. RSV infection usually results in colds with profuse rhinorrhea, but in first-time infections among infants 6 weeks to 6 months old, 25-40% will develop lower respiratory tract illness63. In most areas researched, RSV was responsible for more pneumonia and bronchiolitis than all other microbial pathogens46. Studies suggest that childhood RSV pneumonia and bronchiolitis may result in long term respiratory abnormalities such as abnormal pulmonary function, asthma, and recurrent cough and bronchitis51.

      RSV has also been implicated in sudden infant death syndrome (SIDS), although the nature of the association is undetermined16,46,56,61,69.

      Respiratory syncytial virus belongs to the family Paramyxoviridae and the genus Pneumovirus. It is an enveloped pleomorphic virus ranging from 150-300nm in diameter5,33,56 with a single-stranded RNA genome63.

      RSV may be detected by immunofluorescence, EIA, complement fixation, neutralization, or culture isolation and confirmation21,32,46,49. A variety of cell lines are suitable for RSV cultivation. For primary isolation, Hep-2 or HeLa63 are acceptable cell lines although others such as Vero, LLCMK-2, or CV-l have been used. The virus produces characteristic cytopathic effects of syncytium formation and cell destruction63.
      Materials Required but Not Delivered1. Cell Culture for Isolation of Respiratory Viruses. Each laboratory must maintain viable stocks of cells at appropriate passage-state that will efficiently allow replication of respiratory viruses from processed patient specimens. These cells must be checked periodically for ability to support growth of respiratory viruses. Appropriate cell lines can be obtained from the American Type Culture Collection (ATCC), 12301 Parklawn Drive, Rockville, MD 20852.

      2. Viral Transport Medium which is non-inhibitory to the respiratory viruses and the tissue culture cells used for viral isolation. Hank's balanced salt solution with antibiotics and a protein stabilizer is a suitable medium. Avoid use of animal sera (except precolostral fetal bovine serum) as protein stabilizer to prevent interference from inherent antibody.

      3. Tissue Culture Media such as RPMI or Eagle's Minimum Essential Medium with appropriate amount of precolostral fetal bovine serum.

      4. Sterile Tissue Culture Tubes, Dram Vials, or Multi-well Plates.

      5. Acetone, 99.5%.

      Note: Acetone is hygroscopic and should be kept in tightly stoppered bottles. Presence of moisture in the acetone may result in a hazy appearance on the substrate during fluorescence microscopy.

      6. Acetone Cleaned Glass Slides.

      7. Sterile Pipettes.

      8. Humid Chamber.

      9. Sodium Hypochlorite Solution (0.05%).

      10. No. 1 Coverslips.

      11. Incubator with Rheostat for Temperature Regulation.

      12. Sterile Swabs.

      13. Forceps.

      14. Vials for Collection and Transportation of Specimens.

      15. Fluorescence Microscope with appropriate filter combination for FITC (excitation peak 490 nm, emission peak 520 nm).

      16. Sterile Glass Beads (1-3 mm diameter).

      17. Centrifuge.

      18. Vortex Mixer or Sonicator.

      19. Distilled Water.
      References
      Product Information
      Components
      • Respiratory Viral Screen - (Catalog No. 5007). One (1) 10 ml dropper vial containing monoclonal antibody against: Adenovirus, Influenza A, Influenza B, Parainfluenza 1, Parainfluenza 2, Parainfluenza 3, and Respiratory Syncytial Virus.
      • Normal Mouse Antibody - (Catalog No. 5014). One (1) 10 ml dropper vial containing Normal Mouse Antibody to be used as a negative control.
      • Anti-Mouse IgG/FITC Conjugate - (Catalog No. 5008). One (1) 10 ml dropper vial containing FITC Labeled Goat Anti-Mouse IgG Antibody.
      • PBS - (Catalog No. 5087). One (1) packet of Phosphate Buffered Saline Salts yields 1 liter when dissolved in distilled water. Store in a clean closed container at room temperature.
      • Tween 20 / Sodium Azide Solution (100X) - (Catalog No. 5037) One (1) 10 ml vial containing Polyoxyethylene Sorbitan Monolaurate (Tween 20) and Sodium Azide (NaN3) concentrate to be diluted 1:100 in PBS.
      • Mounting Fluid - (Catalog No. 5013). One (1) 10 ml dropper vial containing Tris Buffer, Glycerin, Fluorescence Enhancer, and Sodium Azide as preservative. Store at room temperature.
      Detection methodFluorescent
      FormatPurified
      Applications
      Key Applications
      • Immunofluorescence
      Biological Information
      HostMouse
      Antibody TypeMonoclonal Antibody
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Usage Statement
      • For in vitro Diagnostic Use
      • CE Mark
      Storage and Shipping Information
      Storage ConditionsWhen stored at 2-8°C, the Respiratory Viral Screen is stable up to the expiration date printed on the kit label.

      During incubation, slides should be protected from light and kept in a humid chamber at the recommended temperature.

      Do not freeze or expose to elevated temperatures.

      A marked decrease in fluorescence may indicate conjugate or antibody deterioration. A positive control should be tested with each specimen to ensure proper functioning of these reagents and proper staining procedure. If after appropriate analysis there is a decrease in staining intensity, discontinue use of the reagents.

      Warnings and Precautions:

      · Sodium Azide (present in the conjugate, respiratory viral screen, PBS solution, and mounting fluid) can form potentially explosive metal azides with plumbing. Flush with large volumes of water to prevent azide build up.

      · Pooling or diluting conjugate or respiratory viral screen antibody may cause erroneous results.

      · Do not use reagents after expiration.

      · Avoid leaving reagents above 2-8°C for extended periods.

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

      · Handle all specimens and materials as potentially infectious. Decontaminate with 0.05% Sodium Hypochlorite (a 1:100 dilution of household bleach).

      · Do not expose reagents to bright light during storage or incubation.

      · 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.

      · Acetone is extremely flammable and harmful if swallowed or inhaled. Keep away from heat, sparks, or flames. Avoid breathing vapor. Use adequate ventilation.

      · Do not mouth pipette reagents.

      · Do not substitute reagents from other manufacturers.

      · Alteration of protocol provided may cause erroneous results.

      · When staining multiple samples on a slide, avoid cross contamination between samples.

      · Normal mouse antibody should be tested with each cell culture isolate. Fluorescence indicates a non-specific reaction and the test is considered invalid.
      Packaging Information
      Material Size10 mL
      Transport Information
      Supplemental Information
      Specifications

      Documentation

      SDS

      Title

      Safety Data Sheet (SDS) 

      References

      Reference overviewPub Med ID
      Expression of the vesicular inhibitory amino acid transporter in pancreatic islet cells: distribution of the transporter within rat islets.
      Steven D Chessler, William T Simonson, Ian R Sweet, Lisa P Hammerle
      Diabetes 51 1763-71 2002

      Show Abstract
      12031963 12031963

      User Guides

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