Improved affinity coupling for antibody microarrays: Engineering of double‐(His)6‐tagged single framework recombinant antibody fragments
Improved affinity coupling for antibody microarrays: Engineering of double‐(His)6‐tagged single framework recombinant antibody fragments
复制标题
改进抗体微阵列的亲和耦合：双 (His)6 标记单框架重组抗体片段的工程
DOI：
10.1002/pmic.200600036
复制
发表时间：
2006
期刊：
影响因子：
3.4
通讯作者：
C. Borrebaeck
中科院分区：
文献类型：
--
作者：
C. Steinhauer;C. Wingren;F. Khan;M. He;M. Taussig;C. Borrebaeck
Antibody‐based microarray is a novel technology with great promise in biomedicine that will provide unique means to perform global proteome analysis. In the process of designing the high‐density antibody microarrays required, several critical key issues have been identified that remain to be resolved. In particular, there is a great need for specific and selective approaches enabling non‐purified probes to be directly purified, orientated and coupled in a generic one‐step procedure directly on the chip. In this study, we report on the successful design of affinity‐tagged human recombinant single‐chain fragment variable antibody fragments for improved affinity coupling in array applications. By replacing the standard single‐histidine (His)6‐tag with a consecutive double‐(His)6‐tag, the binding to Ni2+‐nitrilotriacetic acid‐coated infstrates was significantly improved. Surface plasmon resonance analysis showed a significantly tighter binding with at least a threefold slower dissociation. The improved binding characteristics thus enabled non‐purified probes even in the format of crude expression supernatants to be directly applied thereby eliminating the need for any time‐consuming pre‐purification step(s) prior to the immobilization. While the double‐(His)6‐tag probes were found to be expressed equally well as compared to the single‐(His)6‐tag probes, they displayed better long‐term functional on‐chip stability. Taken together, the results demonstrate the generic potential of double‐(His)6‐tag recombinant antibodies for the facile fabrication of high‐density antibody microarrays.
影响因子：
1.6
作者：
Mohanty, AK;Wiener, MC
通讯作者：
Wiener, MC
蛋白质微阵列：研究癌症的强大工具。
DOI：
--
发表时间：
2002
期刊：
Current opinion in molecular therapeutics.
影响因子：
--
作者：
Sreekumar,Arun;Chinnaiyan,ArulM
通讯作者：
Chinnaiyan,ArulM