NANOELECTRIC CHIP FOR KINASE PROFILING

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 激酶是一组酶，在细胞信号网络中起着至关重要的作用。已有研究表明，某些激酶的过度表达可能导致癌症的发生。许多基于激酶抑制剂的药物已经上市或正在进行癌症治疗的临床开发。对人类基因组的分析已经产生了一个(超过500个)蛋白激酶的大型文库。开发快速高通量筛选蛋白激酶抑制剂的新技术是很重要的。在这里，我们提出了一种使用基于纳米电极阵列(NEA)的纳米电子芯片来开发一种低成本、多路、无标记和超灵敏的激酶图谱技术。利用镶嵌在二氧化硅中的垂直排列的碳纳米管(CNT)来制造单独寻址的NEAs，只露出最末端，然后用适当的多肽底物进一步功能化，并将其封闭在微流控通道中。在激酶和三磷酸腺苷的存在下，相应的肽底物发生磷酸化，引入一个带负电荷的磷酸基团，导致NEA的交流阻抗改变。这种变化的速度与酶的活性和浓度有关。这 这项技术有可能在下午1-100分在~5-10微升的溶液中同时检测多达100个对混合蛋白有反应的多肽底物。这将大大加快用于癌症诊断、治疗监测和药物发现的激酶图谱。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Kinases are a group of enzymes which serve as crucial nodes in cell signaling network. It has been demonstrated that cancers may be resulted from the over expression of certain kinases. Many drugs based on kinase inhibitors are already on the market or in clinical development for cancer treatment. The analysis of the human genome has yielded a large library of (over 500) protein kinases. It is important to develop new technologies for quick high-throughput screening of protein kinase inhibitors. Here we propose the development of a low-cost, multiplex, label-free, and ultrasensitive technique for kinase profiling using a nanoelectronic chip based on nanoelectrode arrays (NEAs). Individually addressed NEAs are fabricated using vertically aligned carbon nanotubes (CNTs) inlaid in SiO2 with only the very end exposed which are further functionalized with proper peptide substrates and enclosed in a microfluidic channel. In the presence of kinases and ATP, the corresponding peptide substrates undergo phosphorylation with the introduction of a negatively charged phosphate group, resulting in a change in the AC impedance of the NEA. The rate of the change is associated with the kinase activity and concentration. This technique could potentially enable the simultaneous detection of up to 100 peptide substrates responding to the mixture of kinases at 1-100 pM in ~5-10 microliter solutions. This will significantly expedite kinase profiling for cancer diagnosis, treatment monitoring, and drug discovery.