The Development of Homogeneous Enzyme Immunoassay Technology for 5-Hydroxytryptophan Detection

• Main Authors: Yi-chi Shih, 史薏琪
• Other Authors: Tina T.-C. Tseng
• Format: Others
• Language: zh-TW
• Published: 2014
• Online Access: http://ndltd.ncl.edu.tw/handle/96646091492157325163

碩士 === 國立臺灣科技大學 === 化學工程系 === 102 === The goal of this study is to develop a less expensive, more convenient, less time-consuming homogenous enzyme immunoassay. Compare to the traditional heterogeneous enzyme immunoassay which requires tedious and time-consuming procedures, expensive detection systems, or involving hazardous chemicals, homogeneous enzyme immunoassay technology utilizes a reporter enzyme, such as glucose-6-phosphate dehydrogenase (G6PDH), which has the characteristics of antibody-induced inhibition and the enzyme will be used as the label of 5-hydroxytryptophan (5-HTP) which is the precursor of serotonin (5-hydroxytryptamine, 5-HT) and plays an important role in neurotransmission and the brain function. Based on the principle of homogeneous enzyme immunoassay, no additional separation process is required during the immunoassay process. The assay signal can be detected using a regular spectrophotometer and the concentration of 5-HTP at nanomolar levels can be detected in a few minutes. In this study, we will first start using protein electrophoresis and Western blot to determine the results of bioconjugation cross-linking and then we will optimize the experimental conditions, such as the activation time for modifying the carboxyl groups of 5-HTP and regent concentrations or the reaction time during the bioconjugation process, finally obtain maximum inhibition of enzyme activity results.