化学
电化学发光
检出限
免疫分析
生物传感器
试剂
分子生物学
细胞
电极
细胞培养
色谱法
抗体
生物物理学
生物化学
遗传学
生物
物理化学
免疫学
作者
Xiaofei Wang,Hongfang Gao,Honglan Qi,Qiang Gao,Chengxiao Zhang
标识
DOI:10.1021/acs.analchem.7b04359
摘要
A simple electrochemiluminescence (ECL) immunoassay based on a proximity hybridization-regulated strategy was developed for highly sensitive and specific detection of cell surface protein and protein-overexpressing cancer cells. A biosensor was fabricated by self-assembling a thiolated capture ss-DNA3 (partially hybridize with ss-DNA1 and ss-DNA2) and blocking with 6-mercapto-1-hexanol on a gold electrode surface. Target protein was simultaneously bound by two ss-DNA-tagged antibody probes (DNA1-Ab1 and DNA2-Ab2), while DNA1 and DNA2 were brought in sufficient proximity and hybridized with capture DNA3 on the surface of the biosensor. After ECL signal reagent Ru(phen)32+ was intercalated into the hybridized ds-DNAs, ECL measurement was performed in the coreactant solution. A “signal on” proximity hybridization-regulated ECL immunoassay for alpha-fetoprotein (AFP) was developed. The ECL intensity increased with the increase of AFP concentration in the range of 0.05–20.0 ng/mL with a detection limit of 6.2 pg/mL. Moreover, the developed ECL method was successfully used to detect AFP-overexpressing cancer cells (MCF-7 cancer cells as model) with a detection limit of 620 cells/mL (∼60 MCF-7 cells in 100 μL of cell suspension) and discriminate AFP expression on different types of the living cell surface. This work for the first time reports a proximity hybridization-regulated ECL immunoassay for the detection of the cell surface protein on a living cell surface with good specificity and sensitivity. This simple, specific, and sensitive strategy is greatly promising for the detection of proteins and specific cells.
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