A filtration-based protein microarray technique.
A filtration-based protein microarray technique.
复制标题
基于过滤的蛋白质微阵列技术。
作者：
Yangqing Xu;G. Bao
蛋白质微阵列是一种研究蛋白质表达谱和蛋白质功能的新兴技术。然而，在目前的设计方法中，蛋白质微阵列的整体性能可能会受到扩散限制动力学的影响。我们开发了一种新的蛋白质微阵列平台，该平台利用蛋白质微阵列印刷在蛋白质可渗透的硝酸纤维素滤膜上的过滤测定。与传统的孵育-振荡法相比，该方法克服了扩散限制。我们证明，这种新技术可以提高10倍的整体反应动力学速率，产生40年的动态范围，并提高检测灵敏度和特异性。此外，使用多层蛋白芯片，至少14个芯片可以同时探测，在一个单一的测定22400个不同的反应。利用过滤法可以更好地发挥藻胆体和量子点等大型荧光染料的优势。通过检测人血浆样本中的癌胚抗原，证实了基于过滤的蛋白质微阵列的潜在临床应用。
Protein microarrays are an emerging technology for studying protein expression profiling and protein functions. However, with the current design approaches, the overall performance of protein microarrays can be compromised by diffusion-limited kinetics. We developed a new protein microarray platform that utilizes a filtration assay with protein microarrays printed on protein-permeable nitrocellulose filter membranes. Compared with protein microarrays assayed with the conventional incubation-shaking method, this new approach overcomes the diffusion limit. We demonstrated that this novel technique can improve the overall reaction kinetic rate by 10-fold, yield a dynamic range of 4 decades, and enhance the assay sensitivity and specificity. Further, using multistacking protein chips, at least 14 chips can be probed simultaneously, with 22400 different reactions in a single assay. The advantages of large fluorescent dyes, such as phycobilisome and quantum dots, can be better exploited using the filtration assay. The potential clinical applications of the filtration-based protein microarrays were demonstrated by detecting carcinoembryonic antigen in human plasma samples.