Preparation and characterization of specific IgY against Helicobacter pylori adhesin A

ZENG Bo; ZHANG Zhongbao; ZHANG Zhi; SUN Lijin; CHEN Yongmei; CAO Qian; QIAN Ying; LI Zaixin

doi:10.13431/j.cnki.immunol.j.20170187

Abstract：Helicobacter pylori (Hp) is known as the main cause of gastritis disorder,but traditional antibiotic therapy against Hp infection is challenged by the increasing tolerance.A high-targeted egg yolk antibody (IgY) has expected to be a mean of passive immunization therapy for Hp infections.Here,a specific IgY targeting to Hp adhesion-A (IgY-HpaA) was prepared and evaluated.In brief,a urease-,oxidase-,catalase-positive strain was isolated from a suspected Hp infected patient,and hpaA partial gene (702 bp) was amplified from the isolated strain using PCR reaction with a specific primer.Recombinant HpaA protein,molecular weight is about 45 KDa,was expressed as soluble protein in the E.coli prokaryotic expression system and then purified from supernatant through Ni2 +-NTA affinity chromatography,and its antigenicity was identified by Western blotting with anti-Hp IgY.Anti-HpaA vaccine was prepared from the emulsion of rHpaA protein with the same volume of Freund's adjuvant.7 days after immunizing chickens,IgY-HpaA was extracted from yolk using water dilution and sodium chloride precipitation.The purity and titer of IgY-HpaA,evaluated by SDS-PAGE and ELISA,reached to 95％ and 1 ∶ 150 000 respectively.A specific recognition of IgY-HpaA to Hp still been detected although diluted by 1 000 times.Remarkably,the growth of Hp can be suppressed significantly by IgY-HpaA at 5 mg/ml in vitro.In summary,the prepared IgY-HpaA possesses a high specific recognition and growth inhibition to Hp in vitro.