DARU Journal of Pharmaceutical Sciences 2008. 16(3):174-181.

A simple and cost-effective method for rapid purification of alkyl hydroperoxide reductase (AhpC) from Helicobacter pylori and its antibody production


Background and the purpose of the study: 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.

Methods and Results: 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.

Major Conclusion: 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.


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