Extension of dynamic range of sensitive nanoparticle-based immunoassays

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• 作者：Heidi Hyyti盲 ; Noora Ristiniemi ; P盲ivi Laitinen ; Timo L枚vgren ; Kim Pettersson
• 关键词：Immunoassay ; Nanoparticle ; Dynamic range ; Desensitization
• 刊名：Analytical Biochemistry
• 出版年：1 February, 2014
• 年：2014
• 卷：446
• 期：Complete
• 页码：82-86
• 全文大小：503 K

文摘

Nanoparticles have successfully been employed in immunometric assays that require high sensitivity. Certain analytes, however, require dynamic ranges (DRs) around a predetermined cut-off value. Here, we have studied the effects that antibody orientation and addition of free solid-phase and detection antibodies have on assay sensitivity and DR in traditional sandwich-type immunoassays. D-dimer and cardiac troponin I (cTnI), both routinely used in critical care testing, were applied as model analytes. The assays were performed in microtitration wells with preimmobilized solid-phase antibody. Inherently fluorescent nanoparticles coated with second antibody were used to detect the analyte. The selection of antibody orientation and addition of free solid-phase or detection antibody, with nanoparticles and calibrator, desensitized the assays and extended the DR. With D-dimer the upper limit of the DR was improved from 50 to 10,000 ng/ml, and with cTnI from 25 to 1000 ng/ml. Regression analysis with the Stago STA Liatest D-dimer assay yielded a slope (95% confidence interval) of 0.09 (0.07-0.11) and a y-intercept of 鈭?.79 (鈭?7.87-2.29) ng/L (n = 65, r = 0.906). Thus it is concluded that Europium(III)-chelate-doped nanoparticles can also be employed in immunoassays that require wide DRs around a certain cut-off limit.