Peoples Michael C, Karnes H Thomas
Department of Pharmaceutics, Virginia Commonwealth University Medical Center, P.O. Box 980533, Richmond, Virginia 23298-0533, USA.
Anal Chem. 2008 May 15;80(10):3853-8. doi: 10.1021/ac800244n. Epub 2008 Apr 10.
A miniaturized system based on microfluidic capillaries is presented for point-of-care testing and clinical assessment. The approach relies on microsyringe pump-generated flow to deliver reagents and immunoaffinity chromatography to isolate the antigen from biological matrixes. Capillary sandwich immunoassays for C-reactive protein (CRP) were demonstrated in human serum and cerebrospinal fluid (CSF), which are relevant matrixes for cardiovascular disease risk and meningitis research, respectively. Capillaries packed with antibody-coated silica beads were used to capture CRP from the matrix and a second, dye-labeled antibody was introduced to form a sandwich complex. An acidic elution buffer dissociated the antibody-antigen complexes, and the labeled antibody was detected with diode laser-induced fluorescence. Four parameter logistic functions and % relative error plots were used to model and assess the data. The calibration ranges for CRP were 0.05-3.0 microg/mL in 1:10 diluted serum and 0.01-30 microg/mL in undiluted CSF. The microfluidic apparatus employed a flow rate of 2 microL/min and a sample injection volume of 250 nL. Since it was not necessary to reach antibody-antigen reaction equilibrium and the assay platform dimensions were minimal, run times were as short as 10 min.
本文介绍了一种基于微流控毛细管的小型化系统，用于即时检测和临床评估。该方法依靠微注射泵产生的流动来输送试剂，并利用免疫亲和色谱从生物基质中分离抗原。在人血清和脑脊液（CSF）中分别演示了用于C反应蛋白（CRP）的毛细管夹心免疫测定，血清和脑脊液分别是心血管疾病风险和脑膜炎研究的相关基质。用包被抗体的硅胶珠填充的毛细管从基质中捕获CRP，并引入第二种染料标记抗体形成夹心复合物。酸性洗脱缓冲液使抗体 - 抗原复合物解离，并用二极管激光诱导荧光检测标记抗体。使用四参数逻辑函数和相对误差百分比图对数据进行建模和评估。CRP在1:10稀释血清中的校准范围为0.05 - 3.0微克/毫升，在未稀释脑脊液中的校准范围为0.01 - 30微克/毫升。微流控装置的流速为2微升/分钟，进样体积为250纳升。由于无需达到抗体 - 抗原反应平衡且检测平台尺寸最小，运行时间短至10分钟。
