Desai Urvee A, Deo Sapna K, Hyland Kenneth V, Poon Michael, Daunert Sylvia
Department of Chemistry, University of Kentucky, Lexington 40506, USA.
Anal Chem. 2002 Aug 1;74(15):3892-8. doi: 10.1021/ac025518v.
This work describes a solid-phase immunoassay for 6-keto-prostaglandin F1alpha, the stable hydrolysis product of prostacyclin (prostaglandin I2). Prostacyclin, a potent vasodilator with antiplatelet and antiproliferative properties is an effective treatment for primary pulmonary hypertension and pulmonary arterial hypertension associated with scleroderma and scleroderma-like syndrome. Levels of 6-keto-prostaglandin F1alpha can be directly correlated with levels of prostacyclin. Therefore, 6-keto-prostaglandin F1alpha, has become the indicator of choice to measure prostacyclin levels. The single-step immunoassay for 6-keto-prostaglandin F1alpha reported here was developed using the bioluminescent protein aequorin as a label. Analyte-label conjugates were constructed by linking the carboxyl group of 6-keto-prostaglandin F1alpha and lysine residues of aequorin by chemical conjugation methods. The binding properties of 6-keto-prostaglandin F1alpha toward its antibody and the bioluminescent properties of aequorin were retained in the conjugate, which was then used to generate a dose-response curve for the analyte in a convenient microtiter plate format. The concentration of 6-keto-prostaglandin F1alpha after extraction from plasma showed good correlation with the concentration of 6-ketoprostaglandin F1alpha obtained without prior extraction of the same plasma sample. This measurement demonstrated that the assay allows the measurement of 6-keto-prostaglandin F1alpha directly in plasma without any pretreatment of the samples, which results in a much simpler method with a faster assay time.
本研究描述了一种用于检测6-酮-前列腺素F1α的固相免疫分析方法，6-酮-前列腺素F1α是前列环素（前列腺素I2）的稳定水解产物。前列环素是一种具有抗血小板和抗增殖特性的强效血管扩张剂，是治疗原发性肺动脉高压以及与硬皮病和硬皮病样综合征相关的肺动脉高压的有效药物。6-酮-前列腺素F1α的水平可直接与前列环素的水平相关联。因此，6-酮-前列腺素F1α已成为测量前列环素水平的首选指标。本文报道的用于检测6-酮-前列腺素F1α的单步免疫分析方法是使用生物发光蛋白水母发光蛋白作为标记物开发的。通过化学偶联方法将6-酮-前列腺素F1α的羧基与水母发光蛋白的赖氨酸残基相连，构建分析物-标记物结合物。6-酮-前列腺素F1α与其抗体的结合特性以及水母发光蛋白的生物发光特性在结合物中得以保留，然后将其用于以方便的微量滴定板形式生成分析物的剂量反应曲线。从血浆中提取后6-酮-前列腺素F1α的浓度与未对同一血浆样品进行预先提取而获得的6-酮-前列腺素F1α的浓度显示出良好的相关性。该测量结果表明，该分析方法允许直接在血浆中测量6-酮-前列腺素F1α，而无需对样品进行任何预处理，这使得该方法更加简单，分析时间更快。
