Sayed Mohammed E, Slusher Aaron L, Ludlow Andrew T
School of Kinesiology, University of Michigan.
School of Kinesiology, University of Michigan;
J Vis Exp. 2019 May 3(147). doi: 10.3791/59550.
The telomere repeat amplification protocol (TRAP) is the most widely used assay to detect telomerase activity within a given a sample. The polymerase chain reaction (PCR)-based method allows for robust measurements of enzyme activity from most cell lysates. The gel-based TRAP with fluorescently labeled primers limits sample throughput, and the ability to detect differences in samples is restricted to two fold or greater changes in enzyme activity. The droplet digital TRAP, ddTRAP, is a highly sensitive approach that has been modified from the traditional TRAP assay, enabling the user to perform a robust analysis on 96 samples per run and obtain absolute quantification of the DNA (telomerase extension products) input within each PCR. Therefore, the newly developed ddTRAP assay overcomes the limitations of the traditional gel-based TRAP assay and provides a more efficient, accurate, and quantitative approach to measuring telomerase activity within laboratory and clinical settings.
端粒重复序列扩增法（TRAP）是检测给定样本中端粒酶活性最广泛使用的检测方法。基于聚合酶链反应（PCR）的方法能够对大多数细胞裂解液中的酶活性进行可靠测量。带有荧光标记引物的基于凝胶的TRAP限制了样本通量，并且检测样本差异的能力仅限于酶活性两倍或更大倍数的变化。液滴数字TRAP（ddTRAP）是一种高度灵敏的方法，它是从传统TRAP检测方法改进而来的，使用户每次运行能够对96个样本进行可靠分析，并获得每个PCR中输入的DNA（端粒酶延伸产物）的绝对定量。因此，新开发的ddTRAP检测方法克服了传统基于凝胶的TRAP检测方法的局限性，为在实验室和临床环境中测量端粒酶活性提供了一种更高效、准确和定量的方法。
