Dispersive solid-phase extraction and immunoassay with internal reference calibration using fatty acid-coated inorganic fluorescent nanoparticles

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• 作者：Consuelo Chá ; fer-Pericá ; s ; Á ; ngel Balaguer ; Á ; ngel Maquieira ; Rosa Puchades ; ^{rpuchades@qim.upv.es}
• 关键词：Nanoparticle ; Immunoassay ; Dispersive SPE ; Fluorescence ; Internal reference ; BSA
• 刊名：Analytical Biochemistry
• 出版年：2013
• 出版时间：1 January, 2013
• 年：2013
• 卷：432
• 期：1
• 页码：31-37
• 全文大小：782 K

文摘

Dispersive solid-phase extraction (dSPE) using fatty acid-coated Eu₂O₃ nanoparticles (NPs) was developed, and a direct immunoassay was carried out employing these NPs as support. Secondary antibodies labeled with fluorophore groups were used as reporters, and the intrinsic optical properties of the Eu₂O₃ NPs were employed as an internal calibration of the detection system. The methodology was optimized for both dSPE¡ªNP amount, sample volume, extraction time, ionic strength, and pH¡ªand immunoassay-immunoreagent concentrations, ionic strength, and incubation time. As proof of concept, the methodology was applied to the bovine serum albumin (BSA)/anti-BSA system, and precision of the method was between 5 % and 17 % with an IC₅₀ of 100 nM. Then, water samples with high saline content (sea water) were assayed to observe the matrix effect, and average recoveries (n = 3) between 78 % and 108 % were obtained, demonstrating the reliability of the developed analytical method. Finally, the simultaneous dSPE-immunoassay methodology was applied to other compounds with very different chemical characteristics such as an oligonucleotide, the antibiotic sulfamerazine, and the pesticide chlorpyriphos. Although the IC₅₀ values for sulfamerazine were approximately 2400 nM, satisfactory standard curves were obtained. However, poor reproducibility and sensitivity results were obtained for the oligonucleotide and chlorpyriphos preliminary assays.