Single-Cell Proteomics

蛋白质组 蛋白质组学 计算生物学 瓶颈 微流控 单细胞分析 核酸 无标记量化 生物 细胞 定量蛋白质组学 纳米技术 计算机科学 生物信息学 基因 遗传学 嵌入式系统 材料科学
作者
Luke Vistain,Savaş Tay
出处
期刊:Trends in Biochemical Sciences [Elsevier BV]
卷期号:46 (8): 661-672 被引量:143
标识
DOI:10.1016/j.tibs.2021.01.013
摘要

Single-cell proteomics holds the potential to create a mechanistic understanding of how the products of many genes interact to produce a cellular phenotype. The recent growth in single-cell profiling has been focused primarily on nucleic acid measurement. There is now a pressing need for more comprehensive protein measurements of single cells, particularly in absolute quantification and multiplexed measurements. Microfluidics is a core enabling technology for absolute quantification. Microfluidic devices can integrate traditional protein quantification technologies into single-cell chambers. There has been rapid growth in multiplexing single-cell protein measurements by incorporating mass spectrometry or DNA sequencing. However, such measurements are still limited by the availability of antibodies. Emerging technologies have the potential to eliminate the need for antibodies, providing an avenue for a minimally biased survey of single-cell expression at the protein level. The inability to make broad, minimally biased measurements of a cell’s proteome stands as a major bottleneck for understanding how gene expression translates into cellular phenotype. Unlike sequencing for nucleic acids, there is no dominant method for making single-cell proteomic measurements. Instead, methods typically focus on either absolute quantification of a small number of proteins or highly multiplexed protein measurements. Advances in microfluidics and output encoding have led to major improvements in both aspects. Here, we review the most recent progress that has enabled hundreds of protein measurements and ultrahigh-sensitivity quantification. We also highlight emerging technologies such as single-cell mass spectrometry that may enable unbiased measurement of cellular proteomes. The inability to make broad, minimally biased measurements of a cell’s proteome stands as a major bottleneck for understanding how gene expression translates into cellular phenotype. Unlike sequencing for nucleic acids, there is no dominant method for making single-cell proteomic measurements. Instead, methods typically focus on either absolute quantification of a small number of proteins or highly multiplexed protein measurements. Advances in microfluidics and output encoding have led to major improvements in both aspects. Here, we review the most recent progress that has enabled hundreds of protein measurements and ultrahigh-sensitivity quantification. We also highlight emerging technologies such as single-cell mass spectrometry that may enable unbiased measurement of cellular proteomes. CyTOF, short for cytometry time of flight, is the commercial name for mass cytometry. This technique applies metal isotope-labeled antibodies to single cells and analyzes them using mass spectrometry. an assay property which defines the range of analyte concentrations that can be quantified. Concentrations below the dynamic range are below the detection limit, and concentrations above the dynamic range result in signal saturation, wherein additional analyte does not increase the measured signal. enzyme-linked immunosorbent assay; one of the most popular versions of a sandwich assay wherein one antibody is used to capture a protein of interest on a surface and a second antibody is used to detect that protein and create a measurable signal. An ELISA uses an enzyme to produce the signal; however, other sandwich assays can use a variety of methods to produce an output. the smallest amount of analyte that generates an unambiguous signal during measurement. Typically, this is calculated as the signal that corresponds to three standard deviations above the measured signal in the absence of analyte. refers to techniques that allow the simultaneous measurement of multiple different biomolecular classes, such as measuring both the transcriptome and proteome from the same sample. a multiplexed assay is capable of measuring several analytes simultaneously. uses a pair of oligonucleotide-conjugated antibodies to translate the detection of a protein into the production of DNA. It has specificity similar to that of an ELSIA (due to requiring two antibodies) but does not require surface binding. the resolution of an assay is the smallest difference between two analyte concentrations that can be reliably detected.
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