Ultra-sensitive analyte detection through kinetic proofreading

目前最先进的免疫吸附分析具有较差的检测限，这使得早期检测病理状态变得更加困难。除了灵敏度低之外，由于非特异性抗体-抗原相互作用，蛋白质的测定也受到低特异性的阻碍。我建议将DNA纳米技术与动力学校对方法相结合，以实现超灵敏、无背景的分析物检测。在这样一篇文章中，短寡核苷酸将聚合，从而将分析物产生的记录转化为可检测的微观结构。聚合反应将由单分子DNA折纸种子引发。因此，可以将检测的灵敏度推向单分子水平。除了有希望的高灵敏度外，高特异度应该通过动态校对步骤来保证。我预计，随着时间的推移，通过摧毁种子来过滤假阳性信号。但是，通过在分析物存在的情况下稳定种子，只有真阳性才能在一定的校对时间内存活下来。我计划从检测DNA开始测试这个概念。特别是像siRNA这样的短寡核苷酸，由于它的长度，目前很难检测到。然后，我将通过估计模型蛋白质的检测极限来将分析扩展到蛋白质。

Current state of the art immunosorbent assays have poor detection limits which makes the early detection of a pathological state more difficult. In addition to low sensitivity, measuring proteins is hampered by low specificity due to unspecific antibody-antigen interactions. I propose to combine DNA nanotechnology with a kinetic proofreading approach for ultrasensitive, background free analyte detection. In such an essay, short oligonucleotides would polymerize and thereby transduce a record created by the analyte into detectable, microscopic structures. The polymerization would be initiated by single molecule DNA origami seeds. Thus, the assay’s sensitivity could be pushed towards the single molecule level. Besides the promising high degree in sensitivity, high specificity should be guaranteed by a kinetic proofreading step. I foresee to filter out false-positive signals by destroying the seeds over time. But, by stabilizing the seed in the presence of an analyte, only true-positives would survive a certain time of proofreading. I plan to test the concept starting with the detection of DNA. Especially short oligonucleotides like siRNA are currently difficult to detect because of its length. Then, I will expand the assay towards proteins by estimating a model protein’s limit of detection.