Digital LAMP in a sample self-digitization (SD) chip.
Digital LAMP in a sample self-digitization (SD) chip.
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DOI：
10.1039/c2lc21247a
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发表时间：
2012-06-21
期刊：
影响因子：
6.1
通讯作者：
Chiu DT
中科院分区：
文献类型：
--
作者：
Gansen A;Herrick AM;Dimov IK;Lee LP;Chiu DT
本文介绍了在样品自数字化（SD）芯片上实现数字环介导DNA扩增（dLAMP）的方法。数字DNA扩增已成为一种有吸引力的技术，以定量DNA的绝对浓度在一个样品。虽然数字聚合酶链式反应仍然是最广泛的应用，但在资源有限的环境中，由于需要热循环和强大的温度控制，其应用受到阻碍。在这种情况下，可以扩增DNA或RNA而无需热循环的等温协议是非常感兴趣的。在这里，我们展示了使用等温数字环介导的DNA扩增在固定液滴阵列中成功扩增单个DNA分子。与大多数（如果不是全部）现有的样品离散方法不同，我们的设计允许芯片上样品体积的自动化，无损数字化。我们证明了在样品体积小于2 μl的情况下精确定量DNA的相对浓度和绝对浓度。在我们的设备中，我们评估了样品自数字化过程中液滴尺寸的均匀性，并验证了尺寸变化足够小，以至于直接计数lamp活性液滴足以进行数据分析。我们预计，我们的SD芯片的简单性和稳健性使其成为一种具有吸引力的廉价和易于操作的DNA扩增设备，例如在护理点设置。
This paper describes the realization of digital loop-mediated DNA amplification (dLAMP) in a sample self-digitization (SD) chip. Digital DNA amplification has become an attractive technique to quantify absolute concentrations of DNA in a sample. While digital polymerase chain reaction is still the most widespread implementation, its use in resource—limited settings is impeded by the need for thermal cycling and robust temperature control. In such situations, isothermal protocols that can amplify DNA or RNA without thermal cycling are of great interest. Here, we showed the successful amplification of single DNA molecules in a stationary droplet array using isothermal digital loop-mediated DNA amplification. Unlike most (if not all) existing methods for sample discretization, our design allows for automated, loss-less digitization of sample volumes on-chip. We demonstrated accurate quantification of relative and absolute DNA concentrations with sample volumes of less than 2 μl. We assessed the homogeneity of droplet size during sample self-digitization in our device, and verified that the size variation was small enough such that straightforward counting of LAMP-active droplets sufficed for data analysis. We anticipate that the simplicity and robustness of our SD chip make it attractive as an inexpensive and easy-to-operate device for DNA amplification, for example in point-of-care settings.
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影响因子：
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