Multiplexed Detection of Small Analytes by Structure-Switching Aptamer-Based Capillary Electrophoresis

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• 作者：Zhenyu Zhu ; Corinne Ravelet ; Sandrine Perrier ; Valrie Guieu ; Batrice Roy ; Christian Perigaud ; Eric Peyrin
• 刊名：Analytical Chemistry
• 出版年：2010
• 出版时间：June 1, 2010
• 年：2010
• 卷：82
• 期：11
• 页码：4613-4620
• 全文大小：246K
• 年卷期：v.82,no.11(June 1, 2010)
• ISSN：1520-6882

文摘

Affinity probe capillary electrophoresis (APCE) assays, combining the separation power of CE with the specificity of interactions occurring between a target and a molecular recognition element (MRE), have become important analytical tools in many application fields. In this report, a rationalized strategy, derived from the structure-switching aptamer concept, is described for the design of a novel APCE mode dedicated to small molecule detection. Two assay configurations were reported. The first one, developed for the single-analyte determination, was based on the use of a cholesteryl-tagged aptamer (Chol-Apt) as the MRE and its fluorescein-labeled complementary strand (CS*) as the tracer (laser-induced fluorescence detection). Under micellar electrokinetic chromatography (MEKC) conditions, free CS* and the hybrid formed with Chol-Apt (duplex*) were efficiently separated (and then quantified) through the specific shift of the electrophoretic mobility of the cholesteryl-tagged species in the presence of a neutral micellar phase. When the target was introduced into the preincubated sample, the hybridized form was destabilized, resulting in a decrease in the duplex* peak area and a concomitant increase in the free CS* peak area. The second format, especially designed for multianalyte sensing, employed dually cholesteryl- and fluorescein-labeled complementary strands (Chol-CS*) of different lengths and unmodified aptamers (Apt). The size-dependent electrophoretic separation of different Chol-CS* forms from each other and from their corresponding duplexes* was also accomplished under MEKC conditions. The simultaneous detection of multiple analytes in a single capillary was performed by monitoring accurately each target-induced duplex-to-complex change. This method could expand significantly the potential of small solute APCE analysis in terms of simplicity, adaptability, generalizability, and high-throughput analysis capability.