Competitive Enzyme ImmunoAssay (ELISA) for the determination of antibodies to Hepatitis A Virus in human plasma and sera.

Regulatery Status: CE
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Product Catalog No: AVAB Pack Size: 96 Wells

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Summary

Hepatitis A continues to be one of the most frequently reported vaccine-preventable diseases in the world, despite the licensure of hepatitis A vaccine in 1995. Widespread vaccination of appropriate susceptible populations would substantially lower disease incidence and potentially eliminate indigenous transmission of hepatitis A virus (HAV) infection.

HAV, a 27-nm RNA agent classified as a picornavirus, can produce either asymptomatic or symptomatic infection in humans after an average incubation period of 28 days (range, 15-50 days). The illness caused by HAV infection typically has an abrupt onset of symptoms that can include fever, malaise, anorexia, nausea, abdominal discomfort, dark urine, and jaundice. The likelihood of having symptoms with HAV infection is related to the person’s age. In children less than 6 years of age, most (70%) infections are asymptomatic; if illness does occur, it is not usually accompanied by jaundice. Among older children and adults, infection is usually symptomatic, with jaundice occurring in greater than 70% of patients. Signs and symptoms usually last less than 2 months, although 10%-15% of symptomatic persons have prolonged or relapsing disease lasting up to 6 months. In infected persons, HAV replicates in the liver, is excreted in bile, and is shed in the stool. Peak infectivity of infected persons occurs during the 2-week period before onset of jaundice or elevation of liver enzymes, when the concentration of virus in stool is highest. The concentration of virus in stool declines after jaundice appears. Children and infants can shed HAV for longer periods than adults, up to several months after the onset of clinical illness. Chronic shedding of HAV in feces does not occur; however, shedding can occur in persons who have relapsing illness. Viremia occurs soon after infection and persists through the period of liver enzyme elevation.

Test Principle

The assay is based on the principle of competition where the antibodies in the sample compete with an anti-HAV specific antibody, labeled with HRP, for a fixed amount of antigen on the solid phase.

A purified and inactivated HAV is coated to the microwells. The patient’s serum/plasma is added to the microwell and antibodies to HAV are captured by the solid phase. After washing, the enzyme conjugate is added and binds to the free HAV antigen, if still present.

The plate is washed to remove unbound conjugate and then the chromogen/substrate is added.

In the presence of peroxidase the colorless substrate is hydrolysed to a coloured end-product, whose optical density may be detected and is inversely proportional to the amount of antibodies to HAV present in the sample.

An additive is added to the sample directly into the well to block interferences able to mask the presence of antibodies, mostly appearing in the follow up of vaccination.

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References
  1. CDC. Summary of notifiable diseases, United States, 1997. MMWR 1998;46:1-87.
  2. CDC. Prevention of hepatitis A through active or passive immunization. Recommendations of the Advisory Committee on Immunization Practices (ACIP). MMWR 1996;45(RR-15).
  3. Krugman S, Giles JP. Viral hepatitis: new light on an old disease. JAMA 1970;212:1019-29.
  4. Hadler SC, Webster HM, Erben JJ, Swanson JE, Maynard JE. Hepatitis A in day-care centers: a communitywide assessment. N Engl J Med 1980;302:1222-7.
  5. Lednar WM, Lemon SM, Kirkpatrick JW, Redfield RR, Fields ML, Kelley PW. Frequency of illness associated with epidemic hepatitis A virus infection in adults. Am J Epidemiol 1985;122:226-33.
  6. Glikson M, Galun E, Oren R, Tur-Kaspa R, Shouval D. Relapsing hepatitis A. Review of 14 cases and literature survey. Medicine 1992;71:14-23.
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