Modeling of homogeneous cloned enzyme donor immunoassay.
Modeling of homogeneous cloned enzyme donor immunoassay.
复制标题
同质克隆酶供体免疫分析的建模。
DOI：
10.1016/j.ab.2004.06.044
复制
发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Andrade,JosephD
中科院分区：
文献类型：
--
作者：
Jeon,SangIl;Yang,Xiaoyun;Andrade,JosephD
用于灵敏检测生物学和临床重要分析物的最广泛使用的分析技术之一是免疫分析。近年来，直接免疫探针允许无标记检测抗体和目标分析物之间的相互作用，已经证明了它们作为快速、简单和高度敏感的方法的能力。克隆酶供体免疫分析（CEDIA）均相分析是基于细菌酶β-半乳糖苷酶，该酶已被基因工程改造成两个无活性片段，酶供体和酶受体。在实验中片段的重新结合形成活性酶，它作用于底物产生有色产物。开发了CEDIA的综合动力学模型，以帮助理解该方法，并促进真正均匀版本的开发，可能适用于试纸型多分析物护理点分析设备（ChemChip）。虽然标准分析涉及两步过程，但我们也选择了单一组合过程的模型，这将更容易应用于ChemChip设备。通过建模仿真，我们得到了产物量和活性酶量的时间过程，在假设参数条件下，两步法和单步法分析物浓度的动态范围分别为10−6-10−7和10−5-10−7M。一种简单的一步免疫分析法具有节省时间和成本、提高动态范围的优点。
One of the most widely used analytical techniques for sensitive detection of biologically and clinically significant analytes is the immunoassay. In recent years direct immunoprobes allowing label-free detection of the interaction between the antibody and the target analyte have proved their capabilities as fast, simple, and nevertheless highly sensitive methods. Cloned enzyme donor immunoassay (CEDIA) homogeneous assay is based on the bacterial enzyme β-galactosidase, which has been genetically engineered into two inactive fragments, enzyme donor and enzyme acceptor. Reassociation of the fragments in the assay forms active enzyme, which acts on substrate to generate a colored product. A comprehensive kinetic model of CEDIA is developed to aid in understanding this method and to facilitate development of a truly homogeneous version, potentially applicable to a dipstick-type multianalyte point of care analytical device (ChemChip). Although the standard assay involves a two-step process, we also chose to model a single-combined process, which would be simpler to apply in a ChemChip device. From the modeling simulation, we obtain the time courses of the amounts of product and active enzyme, from which the dynamic ranges can be obtained as 10−6–10−7and 10−5–10−7M analyte concentration for two-step and single-combined processes under the conditions of the assumed parameters, respectively. A simple one-step immunoassay has the merit of reducing time and cost and has an improved dynamic range.