Electrophoretically mediated microanalysis with small molecules: The Jaffe method for creatinine carried out in a capillary tube
Electrophoretically mediated microanalysis with small molecules: The Jaffe method for creatinine carried out in a capillary tube
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DOI：
10.1002/1522-2683(200107)22:12
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发表时间：
2001-08-01
期刊：
影响因子：
2.9
通讯作者：
Strein, TG
中科院分区：
文献类型：
--
作者：
Kochansky, CJ;Koziol, S;Strein, TG
电泳介导的微分析（EMMA）方法，用于执行两个小分子之间的在线化学，已被表征和优化。“plug-plug”型EMMA方法包括电泳混合和随后在毛细管柱范围内将含肌酸酐样品的纳升塞与碱性苦克酸盐（Jaffe反应）进行反应，毛细管柱作为微反应器。分析是通过压力注射苦味酸塞，然后是含肌酐的样品塞来进行的。然后施加电位以电泳混合两种反应物，孵育时间长达6分钟，以便在应用27 kV分离电位之前进行反应，并在485 nm处进行吸光度检测。使用50 μ m内径（ID）延长光路毛细管（150 μ m路径长度）被发现足以确定人血清中肌酐的升高水平，但不能用于量化正常水平。75 μ ID高敏感细胞（1200 μ路径长度）可定量测定血清中正常和升高的肌酐水平。使用后者对人血清肌酐的校准图在预期的临床范围内，在40 μ m和1.2 mm之间呈线性（r(2) = 0.996），估计检测限为17 μ m（信噪比S/N = 3）。用报道的EMMA方法和公认的临床方法学获得的定量比较结果相关性非常好（斜率= 1.001）。
An electrophoretically mediated microanalysis (EMMA) approach, used to perform online chemistry between two small molecules, has been characterized and optimized. The "plug-plug" type EMMA method involved electrophoretic mixing and subsequent reaction of nanoliter plugs of creatinine-containing samples and alkaline picrate (Jaffe reaction) within the confines of the capillary column, which acts as a microreactor. Analyses were performed by pressure injecting a plug of picrate followed by a plug of the creatinine-containing sample. A potential was then applied to electrophoretically mix the two reactants, and an incubation time of up to 6 min allowed the reaction to proceed prior to the application of a 27 kV separation potential with absorbance detection at 485 nm. The use of a 50 mum inner diameter (ID) extended light path capillary (150 mum pathlength) was found to be adequate for determining elevated levels of creatinine in human blood sera, but could not be used to quantify normal levels. Quantification of both normal and elevated levels of creatinine in sera was possible with a 75 mum ID high-sensitivity cell (1200 mum pathlength). Calibration plots using the latter for creatinine in human blood sera spanned the expected clinical range and were linear between 40 muM and 1.2 mm (r(2) = 0.996) with an estimated limit of detection of 17 mum (signal-to-noise ratio S/N = 3). A quantitative comparison of results obtained with the reported EMMA method and accepted clinical methodology correlated very well (slope = 1.001).