Quackenbush John F, Cassidy Pamela B, Pfeffer Lawrence M, Boucher Kenneth M, Hawkes Jason E, Pfeffer Susan R, Kopelovich Levy, Leachman Sancy A
Huntsman Cancer Institute, University of Utah, Salt Lake City, UT, USA.
Methods Mol Biol. 2014;1102:641-53. doi: 10.1007/978-1-62703-727-3_34.
Intact miRNAs can be isolated from the circulation in significant quantities despite the presence of extremely high levels of RNase activity. The remarkable stability of circulating miRNAs makes them excellent candidates for biomarkers in diagnostic applications as well as therapeutic targets in a variety of disease states including melanoma. Circulating RNA molecules are resistant to degradation by RNases because they are encapsulated in membrane-bound microvesicles. We describe a convenient method for the use of ExoQuick, a proprietary resin developed by Systems Biosciences (Mountain View, CA), whereby microvesicles can be purified under gentle conditions using readily available laboratory equipment. This protocol allows for isolation all microvesicles, regardless of their origin, and provides a convenient method for identifying potential cancer-specific biomarkers from biological fluids including serum and plasma.
尽管存在极高水平的核糖核酸酶活性，但仍可从循环系统中大量分离出完整的微小核糖核酸（miRNA）。循环miRNA具有非凡的稳定性，使其成为诊断应用中生物标志物的理想候选者，以及包括黑色素瘤在内的多种疾病状态下的治疗靶点。循环RNA分子对核糖核酸酶的降解具有抗性，因为它们被包裹在膜结合微泡中。我们描述了一种使用ExoQuick的便捷方法，ExoQuick是由系统生物科学公司（加利福尼亚州山景城）开发的一种专利树脂，通过该方法可以使用现成的实验室设备在温和条件下纯化微泡。该方案允许分离所有微泡，无论其来源如何，并提供了一种从包括血清和血浆在内的生物流体中鉴定潜在癌症特异性生物标志物的便捷方法。
