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Genetic transfer of endothelin converting enzyme activity to CHO‐K1 cells: detection of positive cells by reverse hemolytic plaque assay
Author(s) -
Shiraki Takuma,
Sawamura Tatsuya,
Ikura Tsuyoshi,
Kobayashi Shigeo,
Miwa Soichi,
Masaki Tomoh
Publication year - 1994
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(94)00830-2
Subject(s) - transfection , chinese hamster ovary cell , complementary dna , microbiology and biotechnology , enzyme , cell culture , cloning (programming) , endothelin receptor , biology , enzyme assay , chemistry , cdna library , biochemistry , gene , genetics , receptor , computer science , programming language
We have established a novel method of molecular cloning of endothelin converting enzyme, a key enzyme in the production of a potent vasoconstrictor endothelin‐1, by modification of the reverse hemolytic plaque assay. Also, we demonstrated that a cell line, CHO‐K1, showed no detectable activity of endothelin converting enzyme. This cell line was transfected with a cDNA library of bovine endothelial cells. The modified reverse hemolytic plaque assay was shown to detect even a single CHO‐K 1 cell that was changed to produce mature ET‐1 by transfection. Thus, this novel method is suggested to be useful for the molecular cloning of other secreted antigens and their processing enzyme.