Abstract 1977: Extending the capabilities of a high-parameter immunophenotyping assay with cytoplasmic staining applications for mass cytometry

Abstract

Abstract Mass cytometry, empowered by CyTOF® technology, utilizes monoisotopic metal-tagged antibodies and a high-sensitivity mass cytometer to allow high-dimensional single-cell analysis in complex biological samples. The 30-marker panel Maxpar® Direct™ Immune Profiling Assay™ (Cat. No. 201325) for suspension mass cytometry provides an unprecedented sample-to-answer solution for detecting and analyzing 30 surface markers in a single experiment. With 18 open mass channels for additional biological markers, the Maxpar Direct Assay facilitates panel expansion and enables flexibility for higher multiplexity and applications. Among the potential complementary applications with the Maxpar Direct Assay, intracellular cytokine staining (ICS) is of particular interest as it may be used to assess infiltrating immune cell phenotypes in the tumor microenvironment. However, for the purpose of assessing cell viability in this workflow, the effectiveness of the Cell-ID™ Intercalator-Rh (Cat. No. 201103) that is included in the Maxpar Direct Assay is in question, since cell permeabilization during ICS can potentially damage the DNA-intercalator bondIn this study, we investigated the compatibility of the Cell-ID Intercalator-Rh (103Rh) with intracellular staining. To do this, we stained either human peripheral blood mononuclear cell (PBMC) or whole blood samples (FLDM-400287) with the Maxpar Direct Assay followed by intracellular staining for the detection of expressed cytokines. The intercalator was evaluated for its ability to discriminate live and dead cells when the sample undergoes surface antibody staining. The monoisotopic Cell-ID Cisplatin-194Pt (Cat. No. 201194) was used as the control to provide benchmark measurement of cell viability of the samples. For both sample types, known percentages of heat-killed PBMC were spiked into the samples in order to evaluate the influence of dead cells. We demonstrate that 103Rh provides equivalent functionality as a cell viability indicator during intracellular staining for cytoplasmic proteins compared to Cell-ID Cisplatin-194Pt. This work was designed to support the use of the Maxpar Direct Immune Profiling Assay in combination with additional intracellular markers. Overall, these findings expand the applicability of Cell-ID Intercalator-Rh (103Rh) to processes that involve cytoplasmic staining. For Research Use Only. Not for use in diagnostic procedures. Citation Format: Noah Saederup, Huihui Yao, Michael Cohen, Leslie Fung. Extending the capabilities of a high-parameter immunophenotyping assay with cytoplasmic staining applications for mass cytometry [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1977.