Corstjens Paul L A M, Fidder Herma H, Wiesmeijer Karien C, de Dood Claudia J, Rispens Theo, Wolbink Gert-Jan, Hommes Daniel W, Tanke Hans J
Department of Molecular Cell Biology, Leiden University Medical Center, Einthovenweg 20 Bldg 2, PO Box 9600, 2300 RC, Leiden, The Netherlands.
Anal Bioanal Chem. 2013 Sep;405(23):7367-75. doi: 10.1007/s00216-013-7154-0. Epub 2013 Jul 9.
Monitoring levels of biologicals against tumor necrosis factor (TNF) has been suggested to improve therapeutic outcomes in inflammatory bowel diseases (IBDs). This pilot study describes a rapid lateral flow (LF)-based assay for on-site monitoring of serum trough levels of humanized monoclonal antibody infliximab (IFX). The applied chromatographic method utilizes sequential flows of diluted serum, wash buffer, and an immunoglobulin generic label on LF strips with a Test line comprised of TNF-α. The successive flows permitted enrichment of IFX at the Test line before the label was applied. The label, luminescent upconverting phosphor (UCP) particles coated with protein-A, emits a 550-nm visible light upon excitation with 980-nm infrared light. IFX concentrations were determined through measurement of UCP fluorescence at the Test line. The assay was optimized to detect IFX levels as low as 0.17 μg/mL in serum. For patients with IBD, this limit is appropriate to detect levels associated with loss of response (0.5 μg IFX/mL). The assay was evaluated with clinical samples from patients with Crohn's disease and correlated well within the physiologically relevant range from 0.17 to 10 μg/mL with an IFX-specific ELISA. Performance of the assay was further successfully validated with samples from blood donors, IFX negative IBD patients, and rheumatoid arthritis patients that had developed anti-IFX antibodies. Because of its generic nature, the assay is suited for detecting most therapeutic anti-TNF-α monoclonal antibodies.
监测抗肿瘤坏死因子（TNF）生物制剂的水平已被认为可改善炎症性肠病（IBD）的治疗效果。这项初步研究描述了一种基于快速侧向流动（LF）的检测方法，用于现场监测人源化单克隆抗体英夫利昔单抗（IFX）的血清谷浓度。所应用的色谱方法利用稀释血清、洗涤缓冲液和LF试纸条上的免疫球蛋白通用标记物的顺序流动，试纸条上的检测线由TNF-α组成。连续流动使得在施加标记物之前，IFX在检测线处得以富集。该标记物是涂有蛋白A的发光上转换磷光体（UCP）颗粒，在980 nm红外光激发下发出550 nm可见光。通过测量检测线处UCP的荧光来确定IFX浓度。该检测方法经过优化，可检测血清中低至0.17 μg/mL的IFX水平。对于IBD患者，该限值适用于检测与反应丧失相关的水平（0.5 μg IFX/mL）。该检测方法用来自克罗恩病患者的临床样本进行了评估，在0.17至10 μg/mL的生理相关范围内，与IFX特异性酶联免疫吸附测定法（ELISA）具有良好的相关性。该检测方法还通过来自献血者、IFX阴性IBD患者以及已产生抗IFX抗体的类风湿性关节炎患者的样本进一步成功验证。由于其通用性，该检测方法适用于检测大多数治疗性抗TNF-α单克隆抗体。
