Development Of A Cytokine Chip

细胞因子具有多种调节功能，其中最重要的是调节T细胞分化。分泌的细胞因子谱已被证明可以预测不同的T细胞分化，特别是调节细胞介导性和体液免疫的细胞因子。为了在单个样本中同时测量不同的细胞因子谱，基于对感兴趣的分析物的固相免疫亲和提取，开发了一种基本的细胞因子芯片。重组链霉亲和素斑点阵列通过羰基双咪唑桥被衍生化到硅烷化玻璃载玻片表面。抗细胞因子抗体通过其碳水化合物部分被生物素化，并与Stetavidin结合。样品在分析前用激光染料标记，并与芯片孵化。在通过洗涤去除非反应性物质之后，在扫描激光密度计中读取结合分析物。芯片对所研究的五种细胞因子的灵敏度均为100-150pg/ml。进一步的研究需要对芯片传感器和读数机构进行优化，以进一步提高灵敏度。选择性研究表明，当使用混合细胞因子样本时，该芯片能够具有高度的特异性。新的、选择性更高的芯片的开发为在同一芯片上同时研究许多重要的调控途径打开了大门，如造血、炎症、神经源性调节和伤口愈合。未来的模型预计将包含30-100个配体，使多种分析成为可能。

Cytokines serve a variety of regulatory functions, one of the most important being the regulation of T-cell differentiation. Secreted cytokine profiles have been demonstrated to be predictive of different T-cell differentiation, especially cytokines regulating cell-mediated and humoral immunity. In order to measure different cytokine profiles simultaneously in individual samples, a basic cytokine chip has been developed based on solid-phase immunoaffinity extraction of the analytes of interest. Arrays of recombinant streptavidin spots were derivatized to the surfaces of silanized glass microscope slides via a carbonyldiimidazole bridge. Anti-cytokine antibodies were biotinylated via their carbohydrate moieties and bound to the steptavidin. Samples were labeled with a laser dye prior to analysis and incubated with the chip. Following removal of non-reactive materials by washing, the bound analytes were read in a scanning laser densitometer. Sensitivity of the chip was found to be 100 - 150 pg/ml for all five cytokines studied. Further studies require optimization of the chip sensor and reading mechanism to further increase sensitivity. Selectivity studies indicated that the chip was capable of a high degree of specificity when using mixed cytokine samples. Development of new, increased selectivity chips open the door to studying a number of important regulatory pathways such as hematopoesis, inflammation neurogenic regulation, and wound healing simultaneously on the same chip. Future models are expected to contain 30-100 ligands making multiple analyses possible.