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通过质谱流式细胞术对人呼吸道中分枝杆菌特异性髓样细胞进行表型分析和定量的方案。

Protocol to phenotype and quantify mycobacteria-specific myeloid cells from human airways by mass cytometry.

作者信息

Kiravu Agano, Rozot Virgine, Cruywagen Lauren, Gutschmidt Andrea, DuPlessis Nelita, Nemes Elisa

机构信息

South African Tuberculosis Vaccine Initiative, Institute of Infectious Disease and Molecular Medicine and Division of Immunology, Department of Pathology, University of Cape Town, Cape Town 7925, South Africa.

South African Tuberculosis Vaccine Initiative, Institute of Infectious Disease and Molecular Medicine and Division of Immunology, Department of Pathology, University of Cape Town, Cape Town 7925, South Africa.

出版信息

STAR Protoc. 2024 Dec 20;5(4):103463. doi: 10.1016/j.xpro.2024.103463. Epub 2024 Dec 13.

DOI:10.1016/j.xpro.2024.103463
PMID:39673703
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11699420/
Abstract

Alveolar macrophages and other myeloid cells in the human airways are the primary cell types responding to respiratory pathogens. Here, we present a protocol for in vitro stimulation of cryopreserved human bronchoalveolar lavage (BAL) cells with mycobacterial antigens for phenotyping and quantifying proinflammatory cytokine responses in myeloid cells by mass cytometry. We demonstrate that the measure of markers of myeloid lineage and function is stable after freezing stained cells thereby allowing for batched analyses and/or machine downtime.

摘要

人类气道中的肺泡巨噬细胞和其他髓样细胞是对呼吸道病原体作出反应的主要细胞类型。在此，我们提出了一种用分枝杆菌抗原体外刺激冷冻保存的人支气管肺泡灌洗（BAL）细胞的方案，用于通过质谱流式细胞术对髓样细胞中的促炎细胞因子反应进行表型分析和定量。我们证明，冷冻染色细胞后，髓系谱系和功能标志物的测量是稳定的，从而允许进行批量分析和/或仪器停机。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/c1790d80bbde/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/76157ba1a37b/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/5fd60116d840/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/e0be5c222a09/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/10aa939686d4/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/42d20f2b0035/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/c1790d80bbde/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/76157ba1a37b/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/5fd60116d840/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/e0be5c222a09/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/10aa939686d4/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/42d20f2b0035/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a856/11699420/c1790d80bbde/gr5.jpg

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本文引用的文献

1
Optimising the yield from bronchoalveolar lavage on human participants in infectious disease immunology research.优化感染病免疫学研究中人类参与者支气管肺泡灌洗的产量。
Sci Rep. 2023 May 31;13(1):8859. doi: 10.1038/s41598-023-35723-2.
2
CyTOF for the Masses.流式细胞术大众化。
Front Immunol. 2022 Apr 14;13:815828. doi: 10.3389/fimmu.2022.815828. eCollection 2022.
3
Visualization of Mass Cytometry Signal Background to Enable Optimal Core Panel Customization and Signal Threshold Gating.可视化质谱流式细胞术信号背景以实现最佳核心面板定制和信号阈值门控。
Methods Mol Biol. 2019;1989:35-45. doi: 10.1007/978-1-4939-9454-0_3.
4
Optimization of mass cytometry sample cryopreservation after staining.染色后质谱流式细胞术样本冷冻保存的优化
Cytometry A. 2017 Jan;91(1):48-61. doi: 10.1002/cyto.a.23014. Epub 2016 Oct 31.
5
Multiparameter Phenotyping of Human PBMCs Using Mass Cytometry.使用质谱流式细胞术对人外周血单个核细胞进行多参数表型分析。
Methods Mol Biol. 2015;1343:81-95. doi: 10.1007/978-1-4939-2963-4_7.
6
Normalization of mass cytometry data with bead standards.用标准微球对质谱流式细胞术数据进行标准化处理。
Cytometry A. 2013 May;83(5):483-94. doi: 10.1002/cyto.a.22271. Epub 2013 Mar 19.
7
Mass cytometry: protocol for daily tuning and running cell samples on a CyTOF mass cytometer.质谱流式细胞术：在CyTOF质谱流式细胞仪上进行每日调试和运行细胞样本的方案。
J Vis Exp. 2012 Nov 2(69):e4398. doi: 10.3791/4398.