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Part II. New Tools in Diagnosis |
Exploring cDNA Phage Display for Autoantibody Profiling in the Serum of Multiple Sclerosis PatientsOptimization of the Selection Procedure
C. GOVARTSa,
K. SOMERSa,
R. HUPPERTSb,
P. STINISSENa AND
V. SOMERSa
a Hasselt University, Biomedical Research Institute, and Transnationale Universiteit Limburg, School of Life Sciences, Agoralaan, Diepenbeek, Belgium b Department of Neurology, Academical Hospital Maastricht, Maastricht, the Netherlands
Key Words: serological antigen selection • filamentous phage • cDNA display • multiple sclerosis • autoantibody repertoire • autoimmune disease
Address for correspondence: V. Somers, Hasselt University, Biomedical Research Institute, and Transnationale Universiteit Limburg, School of Life Sciences, Agoralaan, Building A, B-3590 Diepenbeek, Belgium. Voice: 0032-0-11269202; fax: 0032-0-11269209. veerle.somers{at}uhasselt.be
We applied a cDNA phage display method called serological antigen selection (SAS) to identify immunogenic targets that evoke an autoantibody response in the serum of multiple sclerosis (MS) patients. This method involves the display of a cDNA expression library, in this study a MS brain library, on filamentous phage and subsequent selection using patient immunoglobulin G (IgG). To apply the SAS technology for autoantibodies in the serum of MS patients, an optimization was necessary to deplete cDNA products that encode IgG fragments derived from B cells present in the MS brain plaques. We describe a differential screening procedure in which positive selection rounds on MS serum and negative selection rounds on healthy control serum were alternated to optimize the selection procedure. As a result, a substantial decrease of IgG-displaying phage clones was observed after each negative selection round, thereby preventing an overgrowth of IgG-displaying phage clones. Our depletion strategy was therefore successful in preventing the enrichment of IgG-displaying phage clones. This approach will facilitate the identification of possible MS-related antigens.
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