simple and cost-effective method for rapid purification of alkyl hydroperoxide reductase [AhpC] from helicobacterpylori and its antibody production

ABSTRACT

Helicobacter pylori express abundant amounts of AhpC enzyme that functions to reduce organic hydroperoxides [ROOH] into the corresponding non-toxic alcohols [ROH]. This conserved antigen has been earlier described as specific and unique for H. pylori and therefore, both H. pylori AhpC and Anti-AhpC could be useful in the development of serologic and stool antigen tests, to detecting and monitoring H. pylori infection. AhpC may also serves as a potential target for an antimicrobial agent or for vaccine development. The aim of this study was to simplify isolation and purification of the AhpC and production of a highly specific polyclonal antibody against it. In this paper a simple method was used for protein purification and antibody production which avoids both the long term AhpC protein purification procedure and the addition of Freund's adjuvant. One-dimensional preparative gel electrophoresis allows a single and short purification step and the high resolution capacity of this technique leads to a high level of purity of the protein and consequently to a very high specificity of the antibody. Moreover, it avoids contamination by other non-specific proteins which often appear during protein purification by column chromatographic techniques. The present method is simple, rapid and cost-effective and makes it possible to produce antibody for stool antigen enzyme immunoassay in short time and at low cost