preparation of metal-chelating piezoelectric biosensor by hydrogel-coating

• Main Authors: wayne, 王維聰
• Other Authors: Hsiu-Mei Chen
• Format: Others
• Language: zh-TW
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/84024852135103432190

碩士 === 國立臺灣科技大學 === 化學工程系 === 89 === This thesis reported a novel fabrication method, which combined hydrogel-coating and immobilized metal affinity chromatography（IMAC）techniques, to prepare metal-chelating piezoelectric（PZ）biosensity chips for the detection of Schistosoma japonicum glutathion-S-transferase C-terminally fused with polyhistidine tag（SjGST/His）. First, a synthetic chelator, POD/NTA, was prepared by covalently linking nitrilotriacetic acid（NTA）with periodateoxidized dextran（POD）, and firmed by ninhydrin test. Then POD/NTA was covalently immobilized onto a SAM modified PZ chip to prepared a hydrophilic and stable POD/NTA chip. Finally, the prepared chip was used to study the reversible binding of SjGST/His in a continuous-flow injection system. The results showed that the prepared chip was specific for SjGST/His binding after Ni2+ chelating, and regenerable with EDTA with the reusability higher than 10 repetitive assays. The SjGST/His-captured POD/NTA chips showed high immuno-specificity toward anti-SjGST antibody, but showed little or no response toward either anti-BSA antibody or human-IgG. The regenerated POD/NTA chips that had been weekly used to bind SjGST/His were still reusable after 6 weeks proper storage. In addition, the POD/NTA chip was ohowed to be able to correctly detect SjGST/His directly from cell lysate. Finally, the binding kinetics of immobilized Ni2+ with SjGST/His and SjGST were also studied, as well as those of immobilized Co2+.