Studying Glycolytic Oscillations in Individual Yeast Cells by Combining Fluorescence Microscopy with Microfluidics and Optical Tweezers.
Studying Glycolytic Oscillations in Individual Yeast Cells by Combining Fluorescence Microscopy with Microfluidics and Optical Tweezers.
复制标题
通过将荧光显微镜与微流体和光镊相结合来研究单个酵母细胞中的糖酵解振荡。
DOI：
10.1002/cpcb.70
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发表时间：
2019
影响因子：
--
通讯作者：
Gustavsson AK
中科院分区：
文献类型：
--
作者：
Gustavsson AK
在这个单元中，我们提供了一个清晰的阐述方法，用于研究单个细胞的动态响应，使用微流体控制和调整细胞环境，光学镊子精确的细胞定位，和荧光显微镜检测细胞内的反应。本单元着重于单酵母细胞中糖酵解振荡的诱导和研究，但方法可以很容易地调整，以检查其他生物学问题和细胞类型。我们提出了一个分步指南，用于制造微流体装置，用于光学镊子的对准，用于细胞制备，以及用于单细胞中糖酵解振荡的延时成像，包括对常见陷阱的讨论。遵循协议的用户应该能够在长达一小时的过程中检测到清晰的代谢物时间轨迹，这些轨迹指示在快速和可逆的环境调整期间单个细胞中的第二尺度上的动态。John Wiley & Sons，Inc.
In this unit, we provide a clear exposition of the methodology employed to study dynamic responses in individual cells, using microfluidics for controlling and adjusting the cell environment, optical tweezers for precise cell positioning, and fluorescence microscopy for detecting intracellular responses. This unit focuses on the induction and study of glycolytic oscillations in single yeast cells, but the methodology can easily be adjusted to examine other biological questions and cell types. We present a step‐by‐step guide for fabrication of the microfluidic device, for alignment of the optical tweezers, for cell preparation, and for time‐lapse imaging of glycolytic oscillations in single cells, including a discussion of common pitfalls. A user who follows the protocols should be able to detect clear metabolite time traces over the course of up to an hour that are indicative of dynamics on the second scale in individual cells during fast and reversible environmental adjustments. © 2018 by John Wiley & Sons, Inc.