Validation of an enzyme-linked immunosorbent assay for the quantification of human IgG directed against the repeat region of the circumsporozoite protein of the parasite Plasmodium falciparum

Malaria Journal

Methodology

Geert Leroux-Roels¹  Isabelle Desombere¹  Eva Van Braeckel¹  Frederic Clement¹  Vincent Dewar²  Erik Jongert²  Marianne Dewerchin²  Marie-Ange Demoitié²  Pierre Cambron²  Christine Swysen²  Joe Cohen² 

[1] Center for Vaccinology, Ghent University, Ghent, Belgium;GlaxoSmithKline Vaccines, Rixensart, Rue de l’Institut 89, B-1330, Rixensart, Belgium;

关键词: Malaria;    Plasmodium falciparum;    Circumsporozoite protein;    Enzyme-linked immunosorbent assay;    R32LR;    Validation;   

DOI  :  10.1186/1475-2875-11-384

 received in 2012-08-10, accepted in 2012-10-23,  发布年份 2012

来源: Springer

【 摘 要 】

BackgroundSeveral pre-erythrocytic malaria vaccines based on the circumsporozoite protein (CSP) antigen of Plasmodium falciparum are in clinical development. Vaccine immunogenicity is commonly evaluated by the determination of anti-CSP antibody levels using IgG-based assays, but no standard assay is available to allow comparison of the different vaccines.MethodsThe validation of an anti-CSP repeat region enzyme-linked immunosorbent assay (ELISA) is described. This assay is based on the binding of serum antibodies to R32LR, a recombinant protein composed of the repeat region of P. falciparum CSP. In addition to the original recombinant R32LR, an easy to purify recombinant His-tagged R32LR protein has been constructed to be used as solid phase antigen in the assay. Also, hybridoma cell lines have been generated producing human anti-R32LR monoclonal antibodies to be used as a potential inexhaustible source of anti-CSP repeats standard, instead of a reference serum.ResultsThe anti-CSP repeats ELISA was shown to be robust, specific and linear within the analytical range, and adequately fulfilled all validation criteria as defined in the ICH guidelines. Furthermore, the coefficient of variation for repeatability and intermediate precision did not exceed 23%. Non-interference was demonstrated for R32LR-binding sera, and the assay was shown to be stable over time.ConclusionsThis ELISA, specific for antibodies directed against the CSP repeat region, can be used as a standard assay for the determination of humoral immunogenicity in the development of any CSP-based P. falciparum malaria vaccine.

【 授权许可】

Unknown   
© Clement et al.; licensee BioMed Central Ltd. 2012. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.