Pawlak Michael, Schick Eginhard, Bopp Martin A, Schneider Michael J, Oroszlan Peter, Ehrat Markus
Zeptosens AG, Witterswil, Switzerland.
Proteomics. 2002 Apr;2(4):383-93. doi: 10.1002/1615-9861(200204)2:4<383::AID-PROT383>3.0.CO;2-E.
Protein microarrays are considered an enabling technology, which will significantly expand the scope of current protein expression and protein interaction analysis. Current technologies, such as two-dimensional gel electrophoresis (2-DE) in combination with mass spectrometry, allowing the identification of biologically relevant proteins, have a high resolving power, but also considerable limitations. As was demonstrated by Gygi et al. (Proc. Nat. Acad. Sci. USA 2000,97, 9390-9395), most spots in 2-DE, observed from whole cell extracts, are from high abundance proteins, whereas low abundance proteins, such as signaling molecules or kinases, are only poorly represented. Protein microarrays are expected to significantly expedite the discovery of new markers and targets of pharmaceutical interest, and to have the potential for high-throughput applications. Key factors to reach this goal are: high read-out sensitivity for quantification also of low abundance proteins, functional analysis of proteins, short assay analysis times, ease of handling and the ability to integrate a variety of different targets and new assays. Zeptosens has developed a revolutionary new bioanalytical system based on the proprietary planar waveguide technology which allows us to perform multiplexed, quantitative biomolecular interaction analysis with highest sensitivity in a microarray format upon utilizing the specific advantages of the evanescent field fluorescence detection. The analytical system, comprising an ultrasensitive fluorescence reader and microarray chips with integrated microfluidics, enables the user to generate a multitude of high fidelity data in applications such as protein expression profiling or investigating protein-protein interactions. In this paper, the important factors for developing high performance protein microarray systems, especially for targeting low abundant messengers of relevant biological information, will be discussed and the performance of the system will be demonstrated in experimental examples.
蛋白质微阵列被认为是一种使能技术,它将显著扩展当前蛋白质表达和蛋白质相互作用分析的范围。当前的技术,如二维凝胶电泳(2-DE)与质谱联用,虽然能够鉴定生物学相关蛋白质,具有较高的分辨率,但也存在相当大的局限性。正如吉吉等人所证明的(《美国国家科学院院刊》2000年,97卷,9390 - 9395页),从全细胞提取物中观察到的二维凝胶电泳中的大多数斑点来自高丰度蛋白质,而低丰度蛋白质,如信号分子或激酶,仅占少数。蛋白质微阵列有望显著加快新的药物相关标志物和靶点的发现,并具有高通量应用的潜力。实现这一目标的关键因素包括:对低丰度蛋白质定量也具有高读出灵敏度、蛋白质功能分析、短检测分析时间、易于操作以及整合各种不同靶点和新检测方法的能力。泽普托森公司基于专有的平面波导技术开发了一种革命性的新型生物分析系统,该技术使我们能够利用倏逝场荧光检测的特定优势,以微阵列形式进行多重、定量的生物分子相互作用分析,且灵敏度最高。该分析系统包括一个超灵敏荧光读数器和带有集成微流体的微阵列芯片,可让用户在蛋白质表达谱分析或研究蛋白质 - 蛋白质相互作用等应用中生成大量高保真数据。在本文中,将讨论开发高性能蛋白质微阵列系统的重要因素,特别是针对相关生物信息的低丰度信使,并通过实验示例展示该系统的性能。