多色免疫标记临床样本的液质细胞术染色方案。

Mass cytometry immunostaining protocol for multiplexing clinical samples.

机构信息

Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2M9, Canada.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 2M9, Canada; Department of Immunology, University of Toronto, Toronto, ON M5S 1A8, Canada.

出版信息

STAR Protoc. 2022 Aug 22;3(3):101643. doi: 10.1016/j.xpro.2022.101643. eCollection 2022 Sep 16.

DOI:10.1016/j.xpro.2022.101643

PMID:36052346

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC9424627/

Abstract

This is a cytometry by time-of-flight (CyTOF) staining protocol for hematopoietic-derived cells, that leverages live-cell barcoding using receptor-type tyrosine-protein phosphatase C (CD45) antibodies conjugated to metal isotopes in combination with DNA-based palladium barcoding to multiplex up to 40 samples. In this protocol, DNA-based barcoding is performed before surface and intracellular immunostaining, which reduces the batch effects that result from day-to-day variations in staining and instrument sensitivity. This protocol also reduces antibody consumption and eliminates the need for repeated instrument adjustment.

摘要

这是一种基于时间飞行的流式细胞术（CyTOF）染色方案，用于造血细胞，该方案利用受体型酪氨酸蛋白磷酸酶 C（CD45）抗体与金属同位素的连接进行活细胞条形码标记，结合基于 DNA 的钯条形码对多达 40 个样本进行多重分析。在该方案中，在表面和细胞内免疫染色之前进行基于 DNA 的条形码标记，这减少了由于每天的染色和仪器灵敏度变化而导致的批次效应。该方案还减少了抗体的消耗，并消除了对仪器重复调整的需求。