Diagnosis of Helicobacter pylori infection and diseases associated with Helicobacter pylori by Helicobacter pylori outer membrane proteins

Abstract

AIM: To examine the serological response of patients with upper gastrointestinal diseases and Helicobocter pylori (H pylori) infection to two H pylori outer membrane proteins (OMPs) (M_r18000 and M_r26000) acquired by gene recombinant technique, and to determine the diagnostic significance of serological tests derived from these OMPs.

METHODS: Recombinant vectors encoding the two H pylori OMPs were used to transform and express in BL21 (DE3) E.coli. After purification with Ni²⁺-NTA agarose resin, colloid gold kits were prepared with purified recombinant proteins to detect H pylori infection and H pylori-associated diseases by the immunity-marker technology. We selected 150 patients with H pylori infection and digestive symptoms without previous treatment, including chronic gastritis (n = 60), duodenal ulcer (n = 30), gastric ulcer (n = 30), and gastric cancer (n = 30). As controls, 33 H pylori-negative healthy volunteers were also recruited. Serum samples were collected from all subjects, and the antibodies to specific proteins of H pylori were tested with the colloid gold test kits. The sensitivity, specificity and accuracy of the colloid gold tests were evaluated, by using the combination of standard diagnostic methods (¹³C urea breath test and bacteria culture) and classic enzyme-linked immunosorbent assay (ELISA) as reference.

RESULTS: After purification with Ni²⁺-NTA agarose resin, the purity of recombinant fusion proteins was about 95%. The recombinant fusion proteins were recognized by the specific monoclonal antibodies against the two H pylori OMPs, as demonstrated by the ELISA. Of the 150 serum samples from patients infected with H pylori 141 (94.0%) responded positively to the recombinant protein with M_r26000, while the seropositive rates were 95.0%, 96.7%, 96.7% and 90.0% for patients with H pylori-associated chronic gastritis, duodenal ulcer, gastric ulcer, and gastric cancer respectively. The sensitivity, specificity, and accuracy of the colloid gold kit with M_r26000 protein were 94.0%, 97.0%, and 94.5%, respectively. Compared with the classic ELISA, bacteria culture and ¹³C urea breath test results in detecting H pylori-infection, there was no significant difference (P > 0.05). For the colloid gold kit with M_r18000, the seropositive rates were 52.0%, 40.0%, 40.0%, 53.3% and 86.7%, respectively, in H pylori-infected patients, and those with H pylori-associated chronic gastritis, duodenal ulcer, gastric ulcer, and gastric cancer. There was a significant difference (P < 0.05) in seropositivity between patient with gastric cancer (86.7%) and those with other diseases (43.3%).

CONCLUSION: The two colloid gold kits derived from the recombinant OMPs are useful tools either for detecting H pylori infection, or for, predicting H pylori-associated gastric malignancy.

Keywords: $[Keywords]