Evaluation of DNA fragment sizing and quantification by the agilent 2100 bioanalyzer.
Evaluation of DNA fragment sizing and quantification by the agilent 2100 bioanalyzer.
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DOI：
10.1093/clinchem/46.11.1851
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发表时间：
2000-11
影响因子：
9.3
通讯作者：
N. Panaro;Po K i Yuen;T. Sakazume;P. Fortina;L. Kricka;P. Wilding
中科院分区：
文献类型：
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作者：
N. Panaro;Po K i Yuen;T. Sakazume;P. Fortina;L. Kricka;P. Wilding
毛细管电泳（CE）通过向溶液中的样品施加高电压来实现分子种类的有效分离（1）。商业CE装置，可用于略多于十年，已经发现了许多应用（2）（3）（4）（5）（6），但是昂贵（60 000美元）并且需要大量的用户培训和经验。最新进展允许在微芯片设备上进行CE（7）（8）（9）（10）（11）。我们评估了Agilent 2100生物分析仪（Agilent Technologies），它代表了使用该技术的新一代CE仪器。生物分析仪相对便宜（18000美元），操作简单，只需要常规移液和基本的计算机技能。通常，12个核酸样品可以在30分钟内在一次性芯片上进行大小测定和定量。芯片由玻璃制成，包括流体储存器和微通道的互连网络，其必须填充有凝胶染料混合物。每个芯片包含16个威尔斯孔：3个用于装载凝胶-染料混合物，1个用于分子大小梯，12个用于实验样品。核酸通过微通道的运动由一系列电极控制，每个电极独立地连接到公共电源。生物分析仪将数据显示为迁移时间图和计算机生成的虚拟凝胶。传统的CE操作变量[温度、电压、毛细管材料和pH、离子强度和缓冲液（12）的粘度]不能被修改。仪器的价格为18000美元，芯片的价格为12 -18美元/芯片（每个样品1 -1.50美元）。凝胶-染料混合物由线性聚合物和荧光嵌入染料组成。标记物混合物由缓冲液沿着较低和较高分子大小的标记物组成，生物分析仪在测定DNA片段大小时将其用作参考。上部标记也用作计算...的参考。
Capillary electrophoresis (CE) achieves efficient separation of molecular species by the application of high voltages to samples in solution (1). Commercial CE units, available for slightly more than a decade, have found numerous applications (2)(3)(4)(5)(6), but are expensive (∼$60 000) and require substantial user training and experience. Recent advances have allowed CE to be performed on microchip devices (7)(8)(9)(10)(11). We evaluated the Agilent 2100 Bioanalyzer (Agilent Technologies), which represents a new generation of CE instruments that use this technology. The Bioanalyzer is relatively inexpensive (∼$18 000) and is simple to operate, requiring only routine pipetting and basic computer skills. Typically, 12 nucleic acid samples can be sized and quantified on a disposable chip within 30 min. Chips are fabricated from glass and comprise an interconnected network of fluid reservoirs and microchannels, which must be filled with a gel-dye mixture. Each chip contains 16 wells: 3 for loading the gel-dye mixture, 1 for a molecular size ladder, and 12 for experimental samples. The movement of nucleic acids through the microchannels is controlled by a series of electrodes, each of which is independently connected to a common power supply. The Bioanalyzer displays data as both migration-time plots and as computer-generated virtual gels. Traditional CE operating variables [temperature, voltage, capillary material, and pH, ionic strength, and viscosity of buffer (12)] cannot be modified. The instruments costs ∼$18 000, and chips cost ∼$12–18 per chip ($1–1.50 per sample). The gel-dye mixture consists of a linear polymer and a fluorescent, intercalating dye. The marker mixture consists of a buffer along with lower and upper molecular size markers, which the Bioanalyzer uses as references when sizing DNA fragments. The upper marker is also used as a reference for calculating the …