Wang Jiaoning, Ren Jicun
Department of Chemistry, Shanghai Jiaotong University, Shanghai, PR China.
Electrophoresis. 2005 Jun;26(12):2402-8. doi: 10.1002/elps.200410246.
In this paper we have presented a sensitive and rapid immunoassay (IA) method by capillary electrophoresis with an enhanced chemiluminescence detection system (CE-CL) based on the catalytic effects of horseradish peroxidase (HRP) on the luminol-hydrogen peroxide reaction. The conditions for the CL reaction and electrophoresis were systematically investigated using HRP as a model sample. The linear range from 2.5 x 10(-11) to 1.0 x 10(-9) mol/L (R = 0.999), and the detection limit of 1.0 x 10(-12) mol/L (signal-to-noise ratio = 3) for HRP were achieved using para-iodophenol as CL enhancer. The relative standard deviations of the migration time and peak area for 5.0 x 10(-10) mol/L HRP (n = 7) were 0.26 and 4.8%, respectively, using a CE system with a home-built CL detector. Under the optimal condition, the HRP-labeled CA125 antibody (Ab) and the Ab-antigen complex were well separated within 4 min by CE using a high-pH buffer (pH 10.20). The assay was successfully used for quantification of CA125 in human sera from health controls and patients associated with ovarian cancer, and the recoveries of the standard addition experiments were 93-109%. Our primary results demonstrated that IA based on CE-CL detection is a powerful tool for clinical diagnosis combined with these commercial IA kits.
在本文中，我们提出了一种灵敏且快速的免疫分析（IA）方法，即基于辣根过氧化物酶（HRP）对鲁米诺-过氧化氢反应的催化作用，采用增强化学发光检测系统（CE-CL）的毛细管电泳法。以HRP作为模型样品，系统地研究了化学发光反应和电泳的条件。使用对碘苯酚作为化学发光增强剂，实现了HRP的线性范围为2.5×10⁻¹¹至1.0×10⁻⁹ mol/L（R = 0.999），检测限为1.0×10⁻¹² mol/L（信噪比=3）。使用带有自制化学发光检测器的毛细管电泳系统，5.0×10⁻¹⁰ mol/L HRP（n = 7）的迁移时间和峰面积的相对标准偏差分别为0.26%和4.8%。在最佳条件下，使用高pH缓冲液（pH 10.20），通过毛细管电泳在4分钟内可将HRP标记的CA125抗体（Ab）与Ab-抗原复合物良好分离。该分析方法成功用于定量健康对照者和卵巢癌患者血清中的CA125，加标回收实验的回收率为93%-109%。我们的初步结果表明，基于CE-CL检测的免疫分析与这些商用免疫分析试剂盒相结合，是临床诊断的有力工具。
