Hahn-Schickard, Georges-Koehler-Allee 103, 79110 Freiburg, Germany.
Laboratory for MEMS Applications, IMTEK-Department of Microsystems Engineering, University of Freiburg, Georges-Koehler-Allee 103, 79110 Freiburg, Germany.
Biosensors (Basel). 2022 Jun 14;12(6):413. doi: 10.3390/bios12060413.
In this paper, we present the ImmunoDisk, a fully automated sample-to-answer centrifugal microfluidic cartridge, integrating a heterogeneous, wash-free, magnetic- and fluorescent bead-based immunoassay (bound-free phase detection immunoassay/BFPD-IA). The BFPD-IA allows the implementation of a simple fluidic structure, where the assay incubation, bead separation and detection are performed in the same chamber. The system was characterized using a C-reactive protein (CRP) competitive immunoassay. A parametric investigation on air drying of protein-coupled beads for pre-storage at room temperature is presented. The key parameters were buffer composition, drying temperature and duration. A protocol for drying two different types of protein-coupled beads with the same temperature and duration using different drying buffers is presented. The sample-to-answer workflow was demonstrated measuring CRP in 5 µL of human serum, without prior dilution, utilizing only one incubation step, in 20 min turnaround time, in the clinically relevant concentration range of 15-115 mg/L. A reproducibility assessment over three disk batches revealed an average signal coefficient of variation (CV) of 5.8 ± 1.3%. A CRP certified reference material was used for method verification with a concentration CV of 8.6%. Our results encourage future testing of the CRP-ImmunoDisk in clinical studies and its point-of-care implementation in many diagnostic applications.
在本文中，我们提出了 ImmunoDisk，这是一种完全自动化的离心微流控试剂盒，集成了一种异质的、免洗的、基于磁性和荧光珠的免疫分析（结合-游离相检测免疫分析/BFPD-IA）。BFPD-IA 允许实现一个简单的流体结构，其中检测孵育、珠分离和检测都在同一个腔室中进行。该系统使用 C 反应蛋白（CRP）竞争性免疫分析进行了表征。本文介绍了一种针对蛋白质偶联珠在室温下预储存的空气干燥的参数研究。关键参数是缓冲液组成、干燥温度和时间。提出了一种使用不同干燥缓冲液以相同温度和时间干燥两种不同类型蛋白质偶联珠的方案。该方法在临床相关浓度范围（15-115mg/L）内，无需预先稀释，仅使用一个孵育步骤，在 20 分钟的周转时间内，测量 5µL 人血清中的 CRP，实现了从样本到答案的工作流程。对三个磁盘批次的重复性评估显示，信号的平均变异系数（CV）为 5.8±1.3%。使用 CRP 认证参考物质进行方法验证，浓度 CV 为 8.6%。我们的结果鼓励未来在临床研究中对 CRP-ImmunoDisk 进行测试，并在许多诊断应用中实现即时护理实施。
