Sönmez Utku M, Frey Nolan, Minden Jonathan S, LeDuc Philip R
Department of Mechanical Engineering, Carnegie Mellon University.
Department of Biological Sciences, Carnegie Mellon University.
J Vis Exp. 2022 Dec 23(190). doi: 10.3791/64281.
During embryogenesis, coordinated cell movement generates mechanical forces that regulate gene expression and activity. To study this process, tools such as aspiration or coverslip compression have been used to mechanically stimulate whole embryos. These approaches limit experimental design as they are imprecise, require manual handling, and can process only a couple of embryos simultaneously. Microfluidic systems have great potential for automating such experimental tasks while increasing throughput and precision. This article describes a microfluidic system developed to precisely compress whole Drosophila melanogaster (fruit fly) embryos. This system features microchannels with pneumatically actuated deformable sidewalls and enables embryo alignment, immobilization, compression, and post-stimulation collection. By parallelizing these microchannels into seven lanes, steady or dynamic compression patterns can be applied to hundreds of Drosophila embryos simultaneously. Fabricating this system on a glass coverslip facilitates the simultaneous mechanical stimulation and imaging of samples with high-resolution microscopes. Moreover, the utilization of biocompatible materials, like PDMS, and the ability to flow fluid through the system make this device capable of long-term experiments with media-dependent samples. This approach also eliminates the requirement for manual mounting which mechanically stresses samples. Furthermore, the ability to quickly collect samples from the microchannels enables post-stimulation analyses, including -omics assays which require large sample numbers unattainable using traditional mechanical stimulation approaches. The geometry of this system is readily scalable to different biological systems, enabling numerous fields to benefit from the functional features described herein including high sample throughput, mechanical stimulation or immobilization, and automated alignment.
在胚胎发育过程中，协调的细胞运动产生调节基因表达和活性的机械力。为了研究这一过程，人们使用了诸如抽吸或盖玻片压缩等工具对整个胚胎进行机械刺激。这些方法限制了实验设计，因为它们不精确，需要人工操作，并且只能同时处理几个胚胎。微流控系统在自动化此类实验任务的同时提高通量和精度方面具有巨大潜力。本文描述了一种为精确压缩整个黑腹果蝇胚胎而开发的微流控系统。该系统具有带有气动驱动可变形侧壁的微通道，能够实现胚胎对齐、固定、压缩以及刺激后收集。通过将这些微通道并行化为七个通道，可以同时对数百个果蝇胚胎施加稳定或动态的压缩模式。在玻璃盖玻片上制造该系统便于使用高分辨率显微镜对样品进行同步机械刺激和成像。此外，使用生物相容性材料（如聚二甲基硅氧烷）以及使流体在系统中流动的能力，使得该设备能够对依赖培养基的样品进行长期实验。这种方法还消除了对人工安装的需求，人工安装会对样品造成机械应力。此外，能够快速从微通道收集样品使得能够进行刺激后分析，包括组学分析，而传统机械刺激方法无法获得进行此类分析所需的大量样品。该系统的几何形状易于扩展到不同的生物系统，使众多领域能够受益于本文所述的功能特性，包括高样品通量、机械刺激或固定以及自动对齐。
