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Antibody production by injection of living cells expressing non self antigens as cell surface type II transmembrane fusion protein
- Source :
- Journal of Immunological Methods, Vol. 367, no. 1-2, p. 70-77 (2011)
- Publication Year :
- 2011
- Publisher :
- Elsevier BV, 2011.
-
Abstract
- Antigen expression and purification are laborious, time consuming and frequently difficult steps in the process of antibody production. In the present study, we developed a method avoiding these two steps. This method relies on the injection of histocompatible living cells stably expressing the antigen as a cell surface type II transmembrane fusion protein. A vector, nicknamed pCD1-CD134L, was constructed to express the antigen fused at the carboxyterminal end of the human CD134 ligand (CD134L) type II transmembrane protein on the surface of eucaryotic cells. This vector was shown to induce cell surface expression of epitopes from human c-Myc (soluble protein), uterogloblin-related protein 1 (secreted protein) and CD94 (type II transmembrane protein). Using this vector, we developed a method to produce antibodies without antigen production. The flowchart of this method is as follows: (i) cloning of the antigen in the pCD1-CD134L vector; (ii) production of a histocompatible cell line stably expressing the CD134L-antigen fusion protein; (iii) testing for cell surface expression of the fusion protein by targeting the CD134L carrier; and (iv) prime-boost immunisation with living cells expressing the fusion protein. This method was successfully used for production of polyclonal antibodies raised against Ixodes ricinus calreticulin (secreted protein) in mice and for production of monoclonal antibodies raised against an epitope of Vaccinia virus A56 (type I transmembrane protein) protein in rat. The present study is the first to demonstrate the use of a type II transmembrane protein as a carrier for cell surface display of antigens.
- Subjects :
- Vesicle-associated membrane protein 8
Recombinant Fusion Proteins
Immunology
OX40 Ligand
Type II transmembrane protein
Antibody production
Epitope
Retinoblastoma-like protein 1
Mice
Antigen
OX40 Ligand - immunology
Protein A/G
Cell surface display of antigen
Immunology and Allergy
Animals
Immunisation method
Mice, Inbred BALB C
biology
Ixodes
Antibodies, Monoclonal - biosynthesis
Antibodies, Monoclonal
Membrane Proteins
Fusion protein
Molecular biology
Transmembrane protein
Rats
Recombinant Fusion Proteins - immunology
Membrane Proteins - immunology
biology.protein
Female
Protein G
Rabbits
Subjects
Details
- Language :
- English
- Database :
- OpenAIRE
- Journal :
- Journal of Immunological Methods, Vol. 367, no. 1-2, p. 70-77 (2011)
- Accession number :
- edsair.doi.dedup.....cd9f5c9df2649c0db0d6392d4d24b0eb