Abstract
We have established an in vitro immunization method (IVI) of human peripheral blood lymphocytes (PBL) for generating antigen-specific monoclonal antibodies. Until now, we have succeeded in producing monoclonal antibodies specific for a number of soluble antigens. In the present study, we tried to establish an IVI protocol for generating anti-peptide human antibodies. Firstly, we prepared two synthetic peptides from cholera toxin B subunit (pCTB) or rice allergenic protein (pRA). Then we performed IVI of PBL derived from healthy donors by using CTB, RA, and their peptides as the immunized antigens. We measured antibody production levels by a sandwich ELISA. Antigen specificity of produced antibodies were tested by direct ELISA, Western blot and ELISPOT assay. Then we found that anti-CTB antibodies were detected in IVI by using pCTB as antigen. RA-specific antibodies were also generated in IVI against pRA-KLH. In addition, RA-specific B cells were demonstrated to significantly increase by immunization with pRA-KLH, which was evidenced by an ELISPOT assay. These results indicate that anti-peptide antibody can be produced by the IVI method using KLH coupled peptide as sensitizing antigens.
Keywords
- Peripheral Blood Lymphocyte
- Cholera Toxin
- ELISPOT Assay
- Soluble Antigen
- Human Peripheral Blood Lymphocyte
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© 2003 Springer Science+Business Media Dordrecht
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Tamura, T., Katakura, Y., Yamashita, M., Matsumoto, S., Teruya, K., Shirahata, S. (2003). Production of Anti-Peptide Antibody by In Vitro Immunization. In: Yagasaki, K., Miura, Y., Hatori, M., Nomura, Y. (eds) Animal Cell Technology: Basic & Applied Aspects. Animal Cell Technology: Basic & Applied Aspects, vol 13. Springer, Dordrecht. https://doi.org/10.1007/978-94-017-0726-8_45
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DOI: https://doi.org/10.1007/978-94-017-0726-8_45
Publisher Name: Springer, Dordrecht
Print ISBN: 978-90-481-6557-5
Online ISBN: 978-94-017-0726-8
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