| As a serious infection disease, viral hepatitis, is caused by various kinds of hepatitis virus and featured as dangerous infection, complicated route of transmission, broad epidemicity, and high morbidity. Acute infection with hepatitis B virus (HBV) is associated with acute viral hepatitis and begins with general ill-health, loss of appetite, nausea, vomiting, body aches, mild fever, dark urine, and then progresses to development of jaundice. Viral hepatitis is prevalent in the world, especially in China, where there are around 50-70% of the population exposed to the infection of HBV. Hepatitis B (HB) is caused by hepatitis B virus, a kind of hepadnavirus. The genome of Hepatitis B virion is a small circular, partially double stranded DNA, encoding an outer protein shell (envelop, contain HBsAg) and an inner body (core, contain HBcAg, HBeAg, HBV-DNA and DNAP). Viewed with electron microscope, three particles could be identified in human serum containing HBV. They are Dane, spherical and filiform particles. After invading in human liver cells, HBV not only has direct pathopoiesis effect on liver cells but also destroys the viral immune system in those cells through both cell mediated and humoral mediated immunity. Meanwhile, these processes lead to many clinical manifestations. Most patients with acute viral hepatitis could recovery in 6 months. About 10~20% patients are infected with chronic Hepatitis B virus, and eventually suffer hepatic cirrhosis and hepatocellular carcinoma. More than 90% of infected newborns are chronic HBsAg carriers or infected with acute Hepatitis B virus. Hepatitis B vaccines has been used to prevent HBV infection, and the incidence and mortality have been decreased efficiently after inoculation in the epidemic areas over the world.Some HBV variants emerge together with the increasing inoculation of Hepatitis B vaccines and we should be noted to the S gene variants, which might escape from the HBV neutralization antibody. The variants with HBsAg mutation were reported in inoculated subjects and patients grafted with HBV liver from donor who had been received the HB immune globulin(IG) therapy. Some reports indicated that the neutralization antibody induced by common vaccine was not sufficient to recognize these variants. Sequence analysis revealed a single nucleotide mutation in the DNA region coding for the HBsAg"α"epitope, which results in the alteration of amino acid 145 GLy of HBV S gene. The HBsAg"α"epitope might be very important for binding the antibody, and the recognition might be defected due to the mutation in the S variants, so that the variants can still be detected in patients in 5 years. Furthermore, it has been shown that this mutation is replicable and could be stably inherited. Like the wild hepatitis virus, HBsAg"α"145 variants has the ability to infect and transmit, and is associated with the bad prognosis of viral hepatitis. It has been shown that the 145 mutation may attenuate the antigenicity of HBsAg, and subsequently leads to failure of detection by the HBsAg diagnostic reagent. The antibody induced by Hepatitis B vaccines couldn't neutralize the variant. Although most people were inoculated by Hepatitis B vaccines, infection by HBsAg"α"145 variants may become a new public health problem. Therefore, it's urgent to promptly identify whether a subject is infected by such variants with HBsAg"α"epitope mutation.So far, HBV variant could be detected by polymerase chain reaction (PCR) and DNA sequencing technology. But these methods are expensive, time-consuming and difficult in technology when analyzing mass clinical specimens. The 145 nucleotide in HBV S gene mutated with high rate, so it is significant to establish a method to detect such point mutation promptly. In this study, we first cloned HBV S gene, and then prepared HBV variant antigen and its antibodies. Further, the monoclonal and polyclonal antibodies against HBV variant antigen were applied to establish the ELISA-based diagnosis method for HBV. This method was highly specific and sensitive, which was able to resolve the mentioned problems that recessive HBV of persistent infection could not be detected. This method could protect many people from the attack of HBV. The rapid diagnosis assay for HBV mutant strain would be helpful for epidemiological investigation and relationship analysis in term of its heredity and evolution.
|
PDF Full Text Request