Rapid methods for purification of human recombinant interleukin-5 (IL-5) using the anti-murine IL-5 antibody-coupled immunoaffinity column.
A large scale production of human recombinant IL-5 (hIL-5) was performed by way of recombinant DNA technology. In this study, we transfected Chinese hamster ovary cells with pdKCR-hIL-5gene-dhfr plasmid and selected a cell line, with the use of methotrexate, producing large amounts of hIL-5. The recombinant hIL-5 thus obtained induced IgM production of murine B cell leukemia BCL1, and its activity was inhibited by TB13 anti-mouse IL-5 monoclonal antibody. hIL-5 could be purified from the cell-free supernatants of the transfectants with high recoveries by using anti-mouse IL-5 antibody-coupled immunoaffinity column in combination with a gel permeation column chromatography. N terminal amino acid sequence analysis of purified hIL-5 revealed that a single amino-terminal amino acid (isoleucine) is detected and hIL-5 consists of 115 amino acid residues.
Mita
S
S
Department of Biology, Kumamoto University Medical School, Japan.
Hosoya
Y
Y
Kubota
I
I
Nishihara
T
T
Honjo
T
T
Takahashi
T
T
Takatsu
K
K
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D000937:Antigen-Antibody Reactions; D001681:Biological Assay; D015153:Blotting, Western; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D006801:Humans; D015848:Interleukin-5 / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins; D014162:Transfection
D000911:Antibodies, Monoclonal; D015848:Interleukin-5; D011994:Recombinant Proteins