Hepatitis E Virus Antigen ELISA
Cat.No: HEV-AG001
Principle: Two-Step Incubation, Double Antibody “Sandwich”
INTENDED USE
This HEV Ag kit is an enzyme-linked immunosorbent assay for the qualitative detection of hepatitis E viral antigen (HEV Ag) in serum, plasma, or stool samples of human and animal origins. It is intended for use in environmental screening or clinical laboratories for diagnosis and management of infected animals and patients related to infection with hepatitis E virus.
SUMMARY
Hepatitis E virus (HEV) is a non-enveloped, single- stranded RNA virus identified in 1990. Infection with HEV induces acute or sub-clinical liver diseases similar to hepatitis A. HEV infections, endemic and frequently epidemic in developing countries, is seen also in developed countries in a sporadic form with or without a history of traveling to endemic area. The overall case-fatality is 0.5~3%, and much higher (15~25%) among pregnant women.
A hypothesis that HEV infection is a zoonosis was presented in 1995. Then a swine HEV and later an avian HEV were identified and sequenced separately in 1997 and 2001. Since then, HEV infection include anti-HEV, viremia and feces excretion of HEV was seen in a wide variety of animals, i.e., swine, rodents, wild monkeys, deer, cow, goats, dogs and chicken in both the developing and developed countries. A direct testimony was reported that the consumption of uncooked dear meat contaminated with HEV led to acute hepatitis E in human, and HEV genome sequences can be detected in pork livers and biles available in the supermarkets in Japan.
The new HEV antigen ELISA was therefore developed for fast and simple screening of potential reservoirs and secretions of infected animals and humans.
PRINCIPLE OF THE ASSAY
This HEV Ag ELISA kit uses polystyrene microwell strips pre-coated with rabbit anti-HEV antibodies directed against the structural proteins ORF-2 of the native virus. Patient’s or animal´s serum, plasma or PBS dissolved stool sample is added into the microwells.
In case of presence of HEV Ag in the sample, the pre-coated antibodies will be bound to the viral antigen and during the first incubation step, the specific immunocomplex formed is captured on the solid phase. After washing to remove unbound sample, second, monoclonal anti-HEV antibody conjugated to Horseradish Peroxidase (HRP) is added into the wells. During the second incubation step, this antibody will bind to the HEV antigens if they have been captured by anti-HEV antibodies during the first incubation step. The unbound HRP conjugate is removed during washing and Chromogen solutions containing Tetramethylbenzidine (TMB) and urea peroxide are added into the wells. In presence of the antibody-antigen-antibody (HRP) “sandwich” complex, the colorless Chromogens are hydrolyzed by the bound HRP-Conjugate to a blue colored product. The blue color turns yellow after stopping the reaction with sulfuric acid. The amount of color intensity can be measured and is proportional to the amount of antibody captured in the wells, and to the sample respectively. Wells containing samples negative for HEV antigens remain colorless.
COMPONENTS
96 Tests:
- Microwellplate 1plate
- Negative Control 1vial
- Positive control 1vial
- HRP-Conjugate Reagent 1vial
- Stock wash buffer 1bottle
- Chromogen solution A 1vial
- Chromogen solution B 1vial
- Stop solution 1vial
- PLASTIC SEALABLE BAG 1unit
- Plate cover 2 sheets
- Package Inserts 1copy
THE ASSAY PROCEDURE
- Dilute stool sample with PBS for 5 times
- Add sample and Controls each 100µl
- Incubate for 60minutes
- Wash for 5 times
- Add HRP-Conjugate each 100µl
- Incubate for 30minutes
- Wash for 5 times
- Coloring : 50ml A + 50ml B
- Incubate for 30minutes
- Stop the reaction with 50ml stop solution
- Read the absorbance 450nm or 450/630 nm
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