Loebke Christian, Sueltmann Holger, Schmidt Christian, Henjes Frauke, Wiemann Stefan, Poustka Annemarie, Korf Ulrike
Division of Molecular Genome Analysis, German Cancer Research Center, Heidelberg, Germany.
Proteomics. 2007 Feb;7(4):558-564. doi: 10.1002/pmic.200600757.
The advancement of efficient technologies to comply with the needs of systems biology and drug discovery has so far not received adequate attention. A substantial bottleneck for the time-resolved quantitative description of signaling networks is the limited throughput and the inadequate sensitivity of currently established methods. Here, we present an improved protein microarray-based approach towards the sensitive detection of proteins in the fg-range which is based on signal detection in the near-infrared range. The high sensitivity of the assay permits the specific quantification of proteins derived from as little as only 20,000 cells with an error rate of only 5%. The capacity is limited to the analysis of up to 500 different samples per microarray. Protein abundance is determined qualitatively, and quantitatively, if recombinant protein is available. This novel approach was called IPAQ (infrared-based protein arrays with quantitative readout). IPAQ offers a highly sensitive experimental approach superior to the established standard protein quantification technologies, and is suitable for quantitative proteomics. Employing the IPAQ approach, a detailed analysis of activated signaling networks in biopsy samples and of crosstalk between signaling modules as required in drug discovery strategies can easily be performed.
迄今为止，为满足系统生物学和药物发现需求而研发的高效技术尚未得到足够关注。信号网络的时间分辨定量描述存在一个重大瓶颈，即目前已确立的方法通量有限且灵敏度不足。在此，我们提出一种基于蛋白质微阵列的改进方法，用于灵敏检测飞克级别的蛋白质，该方法基于近红外范围内的信号检测。该检测方法的高灵敏度允许对仅来自20,000个细胞的蛋白质进行特异性定量，误差率仅为5%。每个微阵列的分析能力限于最多500个不同样本。如果有重组蛋白，可对蛋白质丰度进行定性和定量测定。这种新方法被称为IPAQ（具有定量读数的基于红外的蛋白质阵列）。IPAQ提供了一种高度灵敏的实验方法，优于已确立的标准蛋白质定量技术，适用于定量蛋白质组学。采用IPAQ方法，可以轻松地对活检样本中激活的信号网络以及药物发现策略中所需的信号模块之间的串扰进行详细分析。
