Detection and Sorting of Extracellular Vesicles and Viruses Using nanoFACS

Q1 Health Professions Current Protocols in Cytometry Pub Date : 2020-12-17 DOI:10.1002/cpcy.81
Aizea Morales-Kastresana, Joshua A. Welsh, Jennifer C. Jones
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引用次数: 6

Abstract

Extracellular vesicles (EVs) are sub-micron-sized membranous spheres secreted by cells. EVs play a functional role as intercellular communicators and are associated with a number of diseases. Research into EVs is an area of growing interest due their many potential uses as therapeutic agents, as diagnostic and theranostic biomarkers, and as regulators of cellular biology. Flow cytometry is a popular method for enumerating and phenotyping EVs, even though the majority of EVs are below the detection sensitivity of most commercially available flow cytometers. Here, we present optimized protocols for EV labeling that increase the signal-to-noise ratio of EVs by removing residual antibody. Protocols for alignment of high-resolution jet-in-air flow cytometers are also provided. Published 2020. U.S. Government.

Basic Protocol 1: Bulk EV staining with CFSE protein binding dye

Basic Protocol 2: Antigen-specific staining of EV markers with fluorochrome-conjugated antibodies

Basic Protocol 3: Astrios EQ instrument setup and sample acquisition

Basic Protocol 4: Counting particles and EVs on Astrios EQ with spike-in reference beads

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利用纳米facs检测和分选细胞外囊泡和病毒
细胞外囊泡(EVs)是细胞分泌的亚微米大小的膜状球体。ev作为细胞间通讯媒介发挥功能作用,并与许多疾病有关。电动汽车的研究是一个越来越受关注的领域,因为它们有许多潜在的用途,如治疗剂、诊断和治疗生物标志物以及细胞生物学的调节剂。流式细胞术是一种流行的枚举和分型ev的方法,尽管大多数ev低于大多数市售流式细胞仪的检测灵敏度。在这里,我们提出了优化的EV标记方案,通过去除残留抗体来提高EV的信噪比。还提供了高分辨率空气射流流式细胞仪的校准方案。2020年出版。美国政府。基本方案1:用CFSE蛋白结合染料进行大量EV染色基本方案2:用荧光染料偶联抗体进行EV标记的抗原特异性染色基本方案3:astros EQ仪器设置和样品获取基本方案4:用参考珠在astros EQ上计数颗粒和EV
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来源期刊
Current Protocols in Cytometry
Current Protocols in Cytometry Health Professions-Medical Laboratory Technology
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期刊介绍: Published in affiliation with the International Society for Advancement of Cytometry, Current Protocols in Cytometry is a "best practices" collection that distills and organizes the absolute latest techniques from the top cytometry labs and specialists worldwide. It is the most complete set of peer-reviewed protocols for flow and image cytometry available.
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Issue Information Detection and Sorting of Extracellular Vesicles and Viruses Using nanoFACS Live Imaging of the Lung Small Particle Fluorescence and Light Scatter Calibration Using FCMPASS Software. Issue Information
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