Abstract
Erythropoietin (EPO) is a growth factor, regulating the proliferation and differentiation of erythroid progenitor cells. In this study, we generated five monoclonal antibodies (mAbs) that reacted specifically with recombinant human EPO (rhEPO). Epitope exclusion and other experiments showed that the mAbs obtained were divided into two groups, differing in recognition sites for rhEPO: group 1 mAbs recognize the N-terminal region of rhEPO, whereas group 2 mAbs seem to recognize a conformation-dependent epitope. Although most of the previously reported anti-EPO antibodies that recognized the N-terminal region of EPO lacked the EPO-neutralizing activity, the group 1 mAbs obtained here had the rhEPO-neutralizing activity. Therefore, the group 1 mAbs may be useful for future study on structure-function relationship of EPO. One of the group 2 mAbs, 5D11A, showed the highest affinity for rhEPO with KD value 0.52 nM and had the highest rhEPO-neutralizing activity. Using this mAb, we developed a reproducible and sensitive enzyme-linked immunosorbent assay for the quantification of bioactive rhEPO.
| Original language | English |
|---|---|
| Pages (from-to) | 181-196 |
| Number of pages | 16 |
| Journal | Journal of Immunoassay and Immunochemistry |
| Volume | 29 |
| Issue number | 2 |
| DOIs | |
| Publication status | Published - Apr 2008 |
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Keywords
- Cell-based potency assay
- Conformation- dependent epitope
- Enzyme-linked immunosorbent assay
- Monoclonal antibodies
- Recombinant human erythropoietin
- Validation
ASJC Scopus subject areas
- Immunology
- Pharmacology
Cite this
Production of novel anti-recombinant human erythropoietin monoclonal antibodies and development of a sensitive enzyme-linked immunosorbent assay for detection of bioactive human erythropoietin. / Yanagihara, Shigehiro; Kori, Yuko; Ishikawa, Rika; Kutsukake, Kazuhiro.
In: Journal of Immunoassay and Immunochemistry, Vol. 29, No. 2, 04.2008, p. 181-196.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Production of novel anti-recombinant human erythropoietin monoclonal antibodies and development of a sensitive enzyme-linked immunosorbent assay for detection of bioactive human erythropoietin
AU - Yanagihara, Shigehiro
AU - Kori, Yuko
AU - Ishikawa, Rika
AU - Kutsukake, Kazuhiro
PY - 2008/4
Y1 - 2008/4
N2 - Erythropoietin (EPO) is a growth factor, regulating the proliferation and differentiation of erythroid progenitor cells. In this study, we generated five monoclonal antibodies (mAbs) that reacted specifically with recombinant human EPO (rhEPO). Epitope exclusion and other experiments showed that the mAbs obtained were divided into two groups, differing in recognition sites for rhEPO: group 1 mAbs recognize the N-terminal region of rhEPO, whereas group 2 mAbs seem to recognize a conformation-dependent epitope. Although most of the previously reported anti-EPO antibodies that recognized the N-terminal region of EPO lacked the EPO-neutralizing activity, the group 1 mAbs obtained here had the rhEPO-neutralizing activity. Therefore, the group 1 mAbs may be useful for future study on structure-function relationship of EPO. One of the group 2 mAbs, 5D11A, showed the highest affinity for rhEPO with KD value 0.52 nM and had the highest rhEPO-neutralizing activity. Using this mAb, we developed a reproducible and sensitive enzyme-linked immunosorbent assay for the quantification of bioactive rhEPO.
AB - Erythropoietin (EPO) is a growth factor, regulating the proliferation and differentiation of erythroid progenitor cells. In this study, we generated five monoclonal antibodies (mAbs) that reacted specifically with recombinant human EPO (rhEPO). Epitope exclusion and other experiments showed that the mAbs obtained were divided into two groups, differing in recognition sites for rhEPO: group 1 mAbs recognize the N-terminal region of rhEPO, whereas group 2 mAbs seem to recognize a conformation-dependent epitope. Although most of the previously reported anti-EPO antibodies that recognized the N-terminal region of EPO lacked the EPO-neutralizing activity, the group 1 mAbs obtained here had the rhEPO-neutralizing activity. Therefore, the group 1 mAbs may be useful for future study on structure-function relationship of EPO. One of the group 2 mAbs, 5D11A, showed the highest affinity for rhEPO with KD value 0.52 nM and had the highest rhEPO-neutralizing activity. Using this mAb, we developed a reproducible and sensitive enzyme-linked immunosorbent assay for the quantification of bioactive rhEPO.
KW - Cell-based potency assay
KW - Conformation- dependent epitope
KW - Enzyme-linked immunosorbent assay
KW - Monoclonal antibodies
KW - Recombinant human erythropoietin
KW - Validation
UR - http://www.scopus.com/inward/record.url?scp=41149123150&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=41149123150&partnerID=8YFLogxK
U2 - 10.1080/15321810801888506
DO - 10.1080/15321810801888506
M3 - Article
C2 - 18360813
AN - SCOPUS:41149123150
VL - 29
SP - 181
EP - 196
JO - Journal of Immunoassay and Immunochemistry
JF - Journal of Immunoassay and Immunochemistry
SN - 1532-1819
IS - 2
ER -