A hybridoma cell line producing the monoclonal antibody against erythropoietin receptor (EpoR) was established using the soluble ectodomain of mouse erythropoietin receptor (sEpoR) as an antigen. The monoclonal antibody termed 1G3 bound to the denatured sEpoR. Epitope mapping with peptide library revealed that 1G3 recognized the amino terminal region including the hexapeptide (positions 6 to 11; LeuProAspProLysPhe). The amino acid sequence in this hexapeptide was identical in mice, rats, and humans, and therefore 1G3 bound to EpoR from all of these sources. Using 1G3, we evaluated sEpoR by a sandwich enzyme-linked immunoassay, and EpoR in the solubilized membrane preparation was detected by Western blotting. The cells expressing EpoR were identified with immunochemical staining. We confirmed the presence of EpoR in a neuronal cell line and PC12 cells, and EpoR was expressed in primary cultured hippocampal neurons.
ARTICLES|
July 15, 1996
Anti-erythropoietin receptor monoclonal antibody: epitope mapping, quantification of the soluble receptor, and detection of the solubilized transmembrane receptor and the receptor-expressing cells
E Morishita,
E Morishita
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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H Narita,
H Narita
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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M Nishida,
M Nishida
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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N Kawashima,
N Kawashima
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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K Yamagishi,
K Yamagishi
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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S Masuda,
S Masuda
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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M Nagao,
M Nagao
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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H Hatta,
H Hatta
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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R Sasaki
R Sasaki
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
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Blood (1996) 88 (2): 465–471.
Citation
E Morishita, H Narita, M Nishida, N Kawashima, K Yamagishi, S Masuda, M Nagao, H Hatta, R Sasaki; Anti-erythropoietin receptor monoclonal antibody: epitope mapping, quantification of the soluble receptor, and detection of the solubilized transmembrane receptor and the receptor-expressing cells. Blood 1996; 88 (2): 465–471. doi: https://doi.org/10.1182/blood.V88.2.465.bloodjournal882465
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