Simplified procedure to recover recombinant antigenized secretory IgA to be used as a vaccine vector.
Induced protection mechanisms at mucosal surfaces involve secretory IgA (SIgA), a complex structure made of polymeric-dimeric IgA (IgA(p/d)) antibody associated with secretory component (SC). SIgA can adhere to M cells of the intestinal and nasal epithelia, are transported across these latter, and are thus available to the immune cells underlying the epithelia. This property makes SIgA suitable as potential mucosal vaccine delivery vector. It remains that production and purification of SIgA is a complex task since IgA(p/d) and SC are naturally synthesized by two different cell types. Furthermore, only IgA(p/d) are capable to associate with SC. Thus, we sought to separate IgA(p/d) and monomeric IgA (IgA(m)) antibodies secreted by hybridoma cells in CELLine bioreactors. To this aim, we connected together two 1-m long columns filled with Sephacryl S-300 beads and placed them under the control of a automatized chromatographic system. In parallel, we produced recombinant antigenized human SC (ra-hSC) in Chinese hamster ovary (CHO) cells adapted to suspension culture in CELLine bioreactors. To avoid intermediate purification of ra-hSC, culture supernatants (SN) containing this latter were combined with purified IgA(p/d), and the recombinant antigenized SIgA (raSIgA) complex was resolved on a 1-m long column filled with Superdex 200 beads. Biochemical characterization based on SDS-PAGE, silver staining, immunodetection and enzyme-linked immunosorbent assay (ELISA) indicates that highly purified raSIgA can be recovered using this simple two-step procedure. Such preparations are currently used to immunize mice to induce mucosal and systemic responses.
Favre
Laurent I
LI
R & D Laboratory, Division of Immunology and Allergy, Centre Hospitalier Universitaire Vaudois, HO 05/1542, Avenue Pierre Decker 4, 1005, Lausanne, Switzerland.
Spertini
François
F
Corthésy
Blaise
B
D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D006801:Humans; D007071:Immunoglobulin A, Secretory / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology
D007071:Immunoglobulin A, Secretory; D011994:Recombinant Proteins