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3123 | LIGHT DIAGNOSTICS™ Influenza A & B DFA, ~50 tests

3123
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      Key ApplicationsFormatHostDetection Methods
      IF FITC M Fluorescent
      Description
      Catalogue Number3123
      Brand Family Chemicon®
      Trade Name
      • LIGHT DIAGNOSTICS
      • Chemicon
      DescriptionLIGHT DIAGNOSTICS™ Influenza A & B DFA, ~50 tests
      OverviewLight Diagnostics™ Influenza A and B DFA Kit is intended for the detection and identification of influenza A and influenza B in respiratory specimens such as throat, nasal and nasopharyngeal swabs, nasopharyngeal aspirates, broncho-alveolar lavages from patients with febrile respiratory illness, following amplification of virus in cell culture. Negative results do not preclude influenza virus infection and should not be used as the sole basis for treatment or other management decisions.

      Performance characteristics for influenza A were established when influenza A/H3 and A/H1 were the predominant influenza A viruses in circulation. When other influenza A viruses are emerging, performance characteristics may vary.

      If infection with a novel influenza A virus is suspected based on current clinical and epidemiological screening criteria recommended by public health authorities, specimens should be collected with appropriate infection control precautions for novel virulent influenza viruses and sent to state or local health departments for testing. Viral culture should not be attempted in these cases unless a Biosafety Level 3 (BSL 3+) facility is available to receive and culture specimens.


      For In Vitro Diagnostic Use.

      Test Principle:

      Light Diagnostics Influenza A and B DFA utilizes a direct immunofluorescent antibody technique for identifying influenza virus in infected cells. The antibody is labeled with fluorescein, which produces an apple-green fluorescence when illuminated with ultraviolet light. The labeled antibody will bind to viral antigen present in the specimen. Unbound reagent is removed by rinsing with phosphate buffered saline (PBS). Antigen-antibody complexes can be visualized by fluorescence microscopy. Uninfected cells stain a dull red due to the presence of Evans Blue in the reagents.

      Summary and Explanation:

      Influenza A and B viruses are members of the Orthomyxoviridae family. They are large enveloped viruses, about 110 nm in diameter, with hemagglutinin (HA) and neuraminidase (NA) projections protruding through the glycoprotein membrane, and containing a segmented, single-stranded RNA (1, 2).

      Both viruses have a high frequency of mutation and cause periodic epidemics of influenza worldwide, which are particularly severe when mutations have resulted in dramatic shifts in the HA or NA structure and the circulating antibodies in a community of people do not recognize the virus strains (2, 3). Being transferred by aerosolized droplets and fomites, along with an incubation period of 1-4 days helps the rapid spread of this highly contagious virus within communities. Specificity in all influenza types is conferred by antigenic differences in two of the major structural proteins - the internal nucleoprotein (NP) and the matrix protein (M) (1, 3).

      Influenza is characterized by tracheobronchitis, pharyngitis, myalgia, fever, headache, and malaise (1, 2, 4). Minimum coryza often distinguishes influenza from other viral respiratory illnesses. Children often experience additional symptoms of gastrointestinal pain, vomiting, myositis, otitis media, conjunctivitis, and croup, and are more likely to show symptoms with influenza B virus rather than influenza A virus. In contrast, the more severe illnesses in adults, resulting in hospitalization, are likely to be caused by influenza A. The most significant complication of influenza is pneumonia, occurring most frequently in the elderly, patients with weakened immune systems, or with chronic kidney disease. Less common complications in adults include Reye's syndrome or other CNS involvement, cardiac symptoms, sinusitis, and otitis media. The increase in deaths from pneumonia during "flu season" is assumed to be due to influenza virus and is used to track influenza epidemics and to check the efficacy of influenza vaccines.

      Since influenza A and B can cause similar symptoms and have similar clinical presentations, all specimens in suspected influenza cases should be tested for both viruses. Identification of the particular virus in useful for epidemiological considerations, because treatment is available for influenza A infections, and to avoid nosocomial spread of the virus. Further, identification of influenza viruses is necessary to differentiate them from other viruses. Mycoplasmas and bacteria can cause similar clinical findings but require different treatment strategies.

      Amantadine and its analog, rimantadine, prevent up to 90% of influenza A infections (not influenza B) and 100% of illnesses if taken prophylactically (1, 5). They can also reduce the duration of illness if taken within the first 2 days of illness.

      Identification and differentiation of influenza A or B in cell culture or patient specimens is usually made using reagents containing antibodies specific for influenza A or influenza B (1). Influenza viruses can be isolated in primary rhesus monkey kidney cells (pRMK), Madin Darby canine kidney (MDCK) cells, and other cell lines, depending on the particular strain of virus (6, 9). The addition of trypsin to the culture fluid at ~2 mg/mL concentration greatly aids in influenza virus isolation in MDCK cells. Since many influenza strains do not cause definable cytopathology, cultures must be tested by hemagglutination (HA) or hemadsorption (HAd ) with guinea pig or chicken erythrocytes to assess viral growth.
      Alternate Names
      • Flu A & Flu B DFA
      Materials Required but Not Delivered· Cell lines susceptible to influenza A and B infection. Each laboratory must maintain viable stocks of cells at appropriate passage level that will efficiently allow replication of influenza A and B from processed patient specimens. These cells must be checked periodically for ability to support growth of influenza A and B. Appropriate cell lines can be obtained from the American Type Culture Collection (ATCC), 12301 Parklawn Drive, Rockville, MD 20852.

      · Viral transport medium, which is non-inhibitory to influenza A and B (Hank's balanced salt solution (HBSS) with antibiotics, or equivalent)

      · Tissue culture media (RPMI or Eagle's minimum essential medium (EMEM) with fetal bovine serum (FBS)), and antibiotics or equivalent

      · Sterile standard tubes, shell vials, or multi-well plates

      · Acetone, reagent grade; stored in glass

      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.

      · Multi-well slides with hydrophobic coating

      · Sterile pipettes

      · Humid chamber

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

      · No. 1 coverslips

      · Incubator, 37° ± 1° C

      · Sterile swabs

      · Forceps

      · Vials for collection and transport of specimens

      · Fluorescence microscope with appropriate filter combination for FITC (excitation peak = 490 nm, emission peak = 520 nm)

      · Sterile glass beads (1 to 3 mm diameter)

      · Centrifuge, capable of 700 to 950 x g, with biohazard buckets and adapters for shell vials

      · Vortex mixer or sonicator

      · Deionized or distilled water
      References
      Product Information
      Components
      • Influenza A Reagent - (Catalog No. 5017). One 2 mL dropper vial containing FITC labeled influenza A antibody, protein stabilizer, Evans Blue and 0.1% sodium azide (preservative).
      • Influenza B Reagent- (Catalog No. 5018). One 2 mL dropper vial containing FITC labeled influenza B antibody, protein stabilizer, Evans blue and 0.1% sodium azide (preservative).
      • Influenza A/B Control Slides - (Catalog No. 5010). Two slides containing one well of influenza A infected cells, one well of influenza B infected cells, and two wells of uninfected cells.
      • Phosphate Buffered Saline - (Catalog No. 5087). One packet of phosphate buffered saline (PBS) salts.
      • Tween 20 / Sodium Azide Solution (100 X) - (Catalog No. 5037). One 10 mL vial containing Tween 20, and sodium azide concentrate.
      • Mounting Fluid - (Catalog No. 5013). One 10 mL dropper vial containing Tris-buffer and glycerin, a fluorescence enhancer, and sodium azide (preservative).
      Detection methodFluorescent
      FormatFITC
      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
      • CE Mark
      • For in vitro Diagnostic Use
      Storage and Shipping Information
      Storage ConditionsWhen stored at 2-8° C, the Light Diagnostics Influenza A and B DFA Kit is stable up to the expiration date printed on the label. Do not freeze or expose to elevated temperatures. Discard any remaining reagents after the kit expiration date. During incubation, slides should be protected from light and kept in a humid chamber at the recommended temperature. A marked decrease in fluorescence may indicate conjugate 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 (NaN3) used as a preservative in the conjugates, PBS/Tween and Mounting Fluid is toxic if ingested. Sodium azide may react with lead or copper plumbing to form highly explosive metal azides (17, 18). Upon disposal, flush with large volumes of water to prevent build-up in plumbing.

      · 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 (bleach).

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

      · Do not mouth-pipette reagents.

      · Avoid contact with Evans Blue (present in conjugates), as it is a potential carcinogen. If skin contact occurs, flush with large volumes of water.

      · Mounting Fluid contains a fluorescence enhancer that may be destructive to mucous membranes. Avoid direct skin or mucous membrane contact. If contact occurs, flush with large volumes of water.

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

      · Do not use reagents beyond expiration date.

      · Do not substitute reagents from other manufacturers.

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

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

      · When staining multiple samples on a slide, avoid cross contamination between samples.
      Packaging Information
      Material Size1 kit
      Transport Information
      Supplemental Information
      Specifications

      Documentation

      MSDS

      Title

      Safety Data Sheet (SDS) 

      References

      Reference overviewPub Med ID
      Phenotypic drug susceptibility assay for influenza virus neuraminidase inhibitors.
      James J McSharry, Ann C McDonough, Betty A Olson, George L Drusano
      Clinical and diagnostic laboratory immunology 11 21-8 2004

      Pokaż streszczenie Pełny tekst artykułu
      14715540 14715540

      User Guides

      Title
      LIGHT DIAGNOSTICS™
      LIGHT DIAGNOSTICS™ INFLUENZA A AND B DFA KIT