Components

Antigen Control Slide - (Catalog No. 5009). Two (2) control slides containing one adenovirus infected (positive) well and one non-infected (negative) well.Anti-Adenovirus / FITC Conjugate - (Catalog No. 5016). One (1) 2 ml dropper vial containing FITC labeled anti-adenovirus antibody in phosphate buffered saline (PBS), pH 7.4 (0.14 M NaCl, 0.008 M Na2HP04 ⋅ 7H20, 0.00l M KH2P04, 0.003 M KCl) with polyoxyethylene sorbitan monolaurate (tween 20), 0.02% Evans blue counterstain, and sodium azide as preservative.Phosphate Buffered Saline - (Catalog No. 5087). One (1) packet of phosphate buffered saline (PBS) salts to yield 1 liter upon reconstitution with 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 of 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.

General description

Light Diagnostics Adenovirus Direct Immunofluorescent Assay (DFA) Kit is intended for in vitro diagnostic use in the qualitative confirmation of adenovirus in inoculated cell cultures.

Test Principle:

The Light Diagnostics Adenovirus DFA Kit utilizes a direct immunofluorescent antibody technique for identifying virus in infected tissue cultures. The antibody is labeled with FITC (fluorescein isothiocyanate) which produces an apple green fluorescence when illuminated with ultraviolet light. The labeled antibody will bind to antigen present in the specimen. The FITC label on the antibody allows the antigen / antibody complex to be visualized by fluorescence microscopy. Positive specimens exhibit apple green cell fluorescence while non-infected cells are stained a dull red due to the presence of Evans Blue in the conjugate diluent. The quality of results will depend on a variety of factors such as the condition of the cells for culture confirmation, the fixative used, and the expertise of the technician performing the test.

Summary and Explanation:

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 adenovirus1. Other adenovirus related lower respiratory illnesses include bronchitis and bronchiolitis2. 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 fever11. 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 infections3.

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 gastroenteritis4,5,6.

Adenovirus has also been linked with intussusception7, acute hemorrhagic cystis8, 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 greater9.

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 diameter2.

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 but it can be done by complement fixation, hemagglutination-inhibition, and neutralization methods10.

Diagnostics Kit

Legal Information

CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany

Linkage

Replaces: 3255

Storage and Stability

When stored at 2-8°C, the Adenovirus kit is stable up to the expiration date printed on the label. 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 (present in the conjugate, diluent, 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 conjugates may cause erroneous results.

⋅ 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 coming in contact with them with the same precautions as for potentially infectious material. Decontaminate with 0.05% sodium hypochlorite (a 1:100 dilution of household bleach).

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

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

⋅ Avoid contact with Evans blue as it is a potential carcinogen. If skin contact occurs, flush with large volumes of water.

⋅ 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.