Development of microchromatography-based immunobiosensor

基于微层析的免疫生物传感器的开发

• 批准号: 12650756
• 负责人: YAMAMOTO Shuichi
• 金额: $ 2.3万
• 依托单位: Yamaguchi University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12650756/
• 关键词: biosensor; chromatography; immunoassay; antigen; antibody; Size-exclusion; ELISA; enzyme immuno assay; 抗原抗体複合体; 免疫反応; 抗体固定化

Various methods have been developed for immunoassay. The most popular method is enzyme-linked immunoadsorbent assay (ELISA), in which antibody is immobilized on the solid phase. In this study, free antigen-antibody complex was measured by using size exclusion chromatography (SEC). For comparison, factors affecting the precision and the reproducibility of 96-well microplate ELISA were examined experimentally. The items tested were antibody concentration, time for antibody immobilization, concentration of contaminant proteins, methods for washing, and etc.Antibody and antigen-protein were successfully separated by micro-SEC (4.6 mm diameter and 30 cm length). However, it was found that quite large antibody-antigen complexes are formed during SEC separation even with monoclonal antibody. The present method is also suitable for characterizing monoclonal antibody properties.

已经开发了各种免疫测定方法。最常用的方法是酶联免疫吸附法（ELISA），将抗体固定在固相上。在本研究中，游离抗原-抗体复合物的测定采用大小排斥色谱法（SEC）。为比较，实验考察了96孔微孔板酶联免疫吸附测定的精密度和重现性的影响因素。检测项目为抗体浓度、抗体固定时间、污染物蛋白浓度、洗涤方法等。采用直径4.6 mm、长30 cm的micro-SEC将抗体与抗原蛋白成功分离。然而，我们发现即使与单克隆抗体分离，SEC也会形成相当大的抗体-抗原复合物。本方法也适用于单克隆抗体性质的表征。

项目成果

S.Yamamoto,E.Miyagawa,: "Dynamic binding perfor mance of large biomolecules such as γ-globulin, virus and virus-like particles on various chromatographic supports,"Progress in Biotechnology,. Vol.16. 81-86 (2000)

S. Yamamoto，E. Miyakawa，“大生物分子（例如 γ-球蛋白、病毒和病毒样颗粒在各种色谱支持物上的动态结合性能）”，Progress in Biotechnology，第 81-86 卷（2000 年）。

宮川英二,山本修一: "ヒトT細胞白血病ウイルスII型特異的モノクロナール抗体によるウイルス抗原精製"化学工学論文集. Vol.27. 186-190 (2001)

Eiji Miyakawa、Shuichi Yamamoto：“使用人 T 细胞白血病病毒 II 型特异性单克隆抗体纯化病毒抗原”，化学工程杂志，第 27 卷，186-190 (2001)。