Rahman Adeeb H, Tordesillas Leticia, Berin M Cecilia
Human Immune Monitoring CoRE, Icahn School of Medicine at Mount Sinai, New York, New York, 10029.
Department of Genetics and Genomics, Icahn School of Medicine at Mount Sinai, New York, New York, 10029.
Cytometry A. 2016 Jun;89(6):601-7. doi: 10.1002/cyto.a.22826. Epub 2016 Apr 6.
The analysis of heterogeneous cell samples by mass cytometry (CyTOF) relies on the assumption that metal labeled antibodies accurately bind to their target antigens. We report a previously unappreciated experimental artifact of non-specific antibody binding by eosinophils during intracellular CyTOF analysis of human whole blood samples. We hypothesized that this non-specific binding results from a charge-based interaction between the metal-labeled antibodies and highly cationic proteins found in eosinophillic granules and found that this non-specific staining artifact could be reduced to background levels with a simple blocking protocol using heparin as a competing anionic protein. This protocol eliminates a potential source of erroneous data interpretation in all experiments involving intracellular staining of human whole blood samples, and allows accurate assessment of dynamic changes in intracellular proteins in eosinophils by CyTOF. © 2016 International Society for Advancement of Cytometry.
通过质谱流式细胞术（CyTOF）对异质细胞样本进行分析，其依据的假设是金属标记抗体能准确结合其靶抗原。我们报告了在对人类全血样本进行细胞内CyTOF分析期间，嗜酸性粒细胞存在非特异性抗体结合这一先前未被认识到的实验假象。我们推测这种非特异性结合是由于金属标记抗体与嗜酸性粒细胞颗粒中发现的高阳离子蛋白之间基于电荷的相互作用所致，并发现使用肝素作为竞争性阴离子蛋白的简单封闭方案可将这种非特异性染色假象降低至背景水平。该方案消除了所有涉及人类全血样本细胞内染色实验中错误数据解读的潜在来源，并允许通过CyTOF准确评估嗜酸性粒细胞内蛋白质的动态变化。© 2016国际细胞计量学促进协会。
