한국수산과학회지 (Korean Journal of Fisheries and Aquatic Sciences)
- 제33권1호
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- Pages.55-59
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- 2000
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- 0374-8111(pISSN)
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- 2287-8815(eISSN)
연어 생식소자극호르몬 II의 Sandwich Enzyme Immunoassay법 개발
Development of a Sandwich Enzyme Immunoassay for Salmon Gonadotropin II.
- KIM Dae-Jung (Taean Marine Hatchery, National Fisheries Research and Development Institute) ;
- HAN Chang-Hee (Depsrment of Biology, Dong-Eui Univenity) ;
- AIDA Katsumi (Depatment of Aquatic Bioscience, Graduate School of Agriculture and Agricultural Life Sciences, The University of Tokyo)
- 발행 : 2000.01.01
초록
무지개 송어의 뇌하수체 및 배양액에 존재하는 GTH II 농도를 측정하기 위해 Avidin- Biotin complex를 이용한 sandwich EIA 계을 개발했다. Protein A sepharose affinity chromatography을 통해서 얻어진 연어 GTH II의 rabbit IgG에 biotinylation시킨 것 (Biotin-salmon GTH II rabbit IgG)을 제2 항체로 사용하였고, Non-Biotin salmon GTH II rabbit IgG는 단지 protein A sepharose affinity chromatography에서 얻어진 IgG를 제 1 항체로 사용하였다. EIA는 sandwich법에 의해서 이루어졌으며, 효소반응 기질로는 TMB(3,3'5,5-tetramethylbenzidine)를 이용했으며, 반응후 450 nm의 흡광도에서 automatic microplate reader로 측정하였다. 그 결과,
A specific and sensitive sandwich enzyme-immunoassay (EIA) using Avidin-Biotin complex was developed for the measurement of GTH II levels in pituitary content and pituitary cell culture medium of the rainbow trout-(Oncorhpchus mykiss). Biotin-salmon GTH II rabbit IgG (sefondary antibody) wai purified by a protein A sepharose affinity chromatography column and that was biotinylated by using Biotin-N-hydroxysuccinimide ofter (BNHS). Non-biotin salmon GTH II rabbit IgG (first antibody) was obtained only through a protein A sepharose affinity chromatography column. The assay was performed by the so-called 'sandwich' method using a microtiter plate, A dose-response curve was obtained between