Preparation Of Anti-UreB And Anti-HpaA Monoclonal Antibodies And Exploration Of Clinical Application

Aim:Helicobacter pylori (Hp) is a gram negative,spiral,microaerophylic bacterium that infects more than 50% of the world population and even 70% in developing country.Developing a non-invasive, convenient and cheap detection method against Hp will contribute a lot to early diagnosis. Nowadays, diagnosis methods of Hp were devided into invasive and non-invasive. Needing no endoscope, non-invasive methods are more appreciated. Of the non-invasive tests now available, serological testing is not accurate among young patients and the C13 urea breath test is expensive. As Hp transmiting is by fecal-oral, it`s possible to detect Hp antigen in stool by ELISA(HpSA). Since approved by FDA to be used in clinic in 1998,the HpSA test has been widely evaluated around the world, with a weighted mean sensitivity and specificity of 90-98%, respectively. The HpSA test is a cheap and useful method for the diagnosis of H pylori infection before and after eradication therapy. It is also an accurate test among all age groups of children.Hp urease B unit(UreB) and Hp adhesin A unit(HpaA) were Hp surface proteins, were conservative in different Hp strains even in Hp coccoids. UreB has a higher proportion in Hp, about 2.5%-5%, and HpaA protein has a higher specificity.Our research aims to prepare the monoclonal antibodies against UreB and HpaA proteins,to detect the specificity of the antibodies, then to establish the sandwich ELISA detection system,and finally to detect the stool samples. The study will contribute to lay a foundation for the clinical diagnosis of Hp.Method:1. Preparation of anti-UreB and anti-HpaA monoclonal antibodies (Mabs). The monoclonal antibodies were prepared with the hybridoma technique and purified with either ammonium sulfate precipitation, caprylic acid- ammonium sulfate method,or protein A affinity chromatograph method.The antibody affinity was compared to determine the optimal purification method.