Self-assembled magnetic matrices for DNA separation chips
Self-assembled magnetic matrices for DNA separation chips
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DOI：
10.1126/science.1068420
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发表时间：
2002-03-22
期刊：
影响因子：
56.9
通讯作者：
Viovy, JL
中科院分区：
文献类型：
--
作者：
Doyle, PS;Bibette, J;Viovy, JL
当一个恒定的、均匀的磁场施加到一个悬浮的超顺磁粒子上，这些粒子被限制在垂直于磁场的薄间隙中，这些粒子自组织成一个固定的、准规则的列阵（图1A）(1)。在现场关闭后，阵列立即返回液体悬浮液。通过改变细胞大小和颗粒浓度，柱间距可以从亚微米调整到约100米。孔径在小于5%的范围内可重现，并且与磁场强度无关，只要它能在分离过程中保持阵列的位置（通常为10 mT，这是用普通永磁体可以实现的）。我们在这里使用平均粒子间距离为5.7 m的自组装阵列，在软光刻制备的微通道设备中分离大的双链DNA[在手工细胞中的初步演示在(2)中报道]。目前使用脉冲场琼脂糖凝胶电泳（PFGE）在12至24小时内进行这种分离。用聚合物溶液对短DNA进行毛细管电泳分析是非常成功的，但大于几个kbp的DNA会导致电流体动力学不稳定(3)。微制造障碍阵列(4)、微流控单分子计数装置或熵阱(5)提供了一种有吸引力的凝胶替代方案，但需要高分辨率微光刻和非常窄的通道（在微米或亚微米范围内），这在成本和“现实生活”样品的稳健性方面提出了挑战。图1C显示了完整DNA[48.5千碱基对（kbp）]和Xho I酶切产生的片段（15和33.5千碱基对）在4.8、7.0和10.0 V/cm电场下的电泳图。三个与单一大小的DNA相关的峰，通过峰强度和单分散样品的穗在10到15分钟内被清楚地识别出来。每次运行之间更换基质，运行间的重现性优于6%。-DNA连接体（包含48.5、97和145.5 kbp的分子和少量更大的分子）
When a constant, homogenous magnetic field is applied to a suspension of superparamagnetic particles confined in a thin gap perpendicular to the field, the particles self-organize into a fixed, quasi-regular array of columns (Fig. 1A)(1). The array returns to a liquid suspension immediately upon field switch-off. The column spacing can be tuned from submicrometer to about 100 m, by varying cell size and particle concentration. The pore size is reproducible within better than 5% and independent of the strength of the magnetic field, so long as it can hold the array in place during separation (typically 10 mT, which is achievable with ordinary permanent magnets).We used here a self-assembled array with an average interparticle distance of 5.7 m to separate large duplex DNA in a microchannel device prepared by soft lithography [preliminary demonstrations in a handmade cell were reported in (2)]. Such separations are currently performed in 12 to 24 hours with pulsed-field agarose gel electrophoresis (PFGE). Analysis of short DNA by capillary electrophoresis with polymer solutions was very successful, but DNA larger than a few kbp led to electrohydrodynamic instabilities (3). Microfabricated arrays of obstacles (4), microfluidic single-molecule counting devices, or entropic traps (5) provide an appealing alternative to gels but require high-resolution microlithography and very narrow channels (in the micrometer or submicrometer range), raising challenges in terms of cost and robustness to “real-life” samples. Figure 1C shows electrophoregrams for intact DNA [48.5 kilobase pairs (kbp)] and fragments (15 and 33.5 kbp) created by Xho I digestion, at fields of 4.8, 7.0, and 10.0 V/cm. Three peaks associated with a single size of DNA, by peak intensity and spiking with monodisperse samples are clearly identified in only 10 to 15 min. The run-to-run reproducibility, with replacement of the matrix between each run, was better than 6%.-DNA concatemers (containing molecules of 48.5, 97, and 145.5 kbp and small amounts of larger