Mei Henrik E, Leipold Michael D, Maecker Holden T
The Human Immune Monitoring Center (HIMC), Institute for Immunity, Transplantation and Infection, Stanford University School of Medicine, Stanford, California, 94305.
Cytometry A. 2016 Mar;89(3):292-300. doi: 10.1002/cyto.a.22778. Epub 2015 Sep 10.
Mass cytometry has overcome limitations of fluorescent single cell cytometry by allowing for the measurement of up to currently ∼40 different parameters on a single cell level. However, the cellular proteome comprises many more potential analytes, and current mass cytometry instrumentation allows for theoretically up to 121 different mass detection channels. The labeling of specific probes with appropriate metal ions is a significant hurdle for exploiting more of mass cytometry's analytical capacity. To this end, we here describe the labeling of antibody with natural abundance or isotopically purified platinum as formulated in cisplatin and circumventing the use of chelator-loaded polymers. We confirm the utility of cisplatin-antibody-conjugates for surface, intracellular, and phosphoepitope-specific immunophenotyping, as well as for application in cell surface CD45-based barcoding. Cisplatin-labeling of antibody increases the analytical capacity of the CyTOF(®) platform by two channels based on available reagents, and has the potential to add a total of six channels for detection of specific probes, thus helping to better extend the analytical mass range of mass cytometers.
质谱流式细胞术克服了荧光单细胞流式细胞术的局限性,能够在单细胞水平上测量多达约40种不同参数。然而,细胞蛋白质组包含更多潜在的分析物,目前的质谱流式细胞术仪器理论上允许多达121个不同的质量检测通道。用适当的金属离子标记特定探针是充分利用质谱流式细胞术分析能力的一个重大障碍。为此,我们在此描述了用天然丰度或同位素纯化的铂(如顺铂中所含)标记抗体,并避免使用负载螯合剂的聚合物。我们证实了顺铂-抗体缀合物在表面、细胞内和磷酸表位特异性免疫表型分析中的效用,以及在基于细胞表面CD45的条形码技术中的应用。基于现有试剂,抗体的顺铂标记使CyTOF(®)平台的分析能力增加了两个通道,并且有可能总共增加六个通道用于检测特定探针,从而有助于更好地扩展质谱细胞仪的分析质量范围。