Nanofluidic chips for cryo-EM structure determination from picoliter sample volumes.
Nanofluidic chips for cryo-EM structure determination from picoliter sample volumes.
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DOI：
10.7554/elife.72629
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发表时间：
2022-01-21
期刊：
影响因子：
7.7
通讯作者：
Jakobi AJ
中科院分区：
文献类型：
--
作者：
Huber ST;Sarajlic E;Huijink R;Weis F;Evers WH;Jakobi AJ
低温电子显微镜已成为生物大分子结构测定的重要工具。在实践中，可靠地制备具有均匀冰厚度的样品的困难仍然是常规高分辨率成像的障碍，并限制了该技术的当前吞吐量。我们表明，纳米流体样品支持具有明确的几何形状，可用于制备冷冻EM标本可重复的冰厚度从皮升样品体积。样品溶液包含在电子透明的纳米通道中，该通道提供均匀的厚度梯度，无需进一步优化，并消除了潜在的破坏性空气-水界面。我们证明了使用三种标准蛋白质样本进行高分辨率结构测定的可能性。纳米制造的样品支撑件具有使冷冻EM工作流程自动化的潜力，并探索冷冻EM应用的新前沿，例如时间分辨成像和高通量筛选。
Cryogenic electron microscopy has become an essential tool for structure determination of biological macromolecules. In practice, the difficulty to reliably prepare samples with uniform ice thickness still represents a barrier for routine high-resolution imaging and limits the current throughput of the technique. We show that a nanofluidic sample support with well-defined geometry can be used to prepare cryo-EM specimens with reproducible ice thickness from picoliter sample volumes. The sample solution is contained in electron-transparent nanochannels that provide uniform thickness gradients without further optimisation and eliminate the potentially destructive air-water interface. We demonstrate the possibility to perform high-resolution structure determination with three standard protein specimens. Nanofabricated sample supports bear potential to automate the cryo-EM workflow, and to explore new frontiers for cryo-EM applications such as time-resolved imaging and high-throughput screening.