Abstract 629: Benefit of using a spectral flow analyzer for the analysis of immune cell populations in tumors

Abstract

Abstract Flow cytometry is a widely applied approach for exploratory immune profiling and enables the identification of individual cells in a cell population, such as a tumor cell suspension, by staining cell markers with antibodies conjugated with a fluorophore. The markers allow for the differentiation of a large variety of immune cell populations, their change in frequency, and activation status upon treatment. Conventional flow cytometers, which use bandpass filters and need conventional compensation matrices, are limited by the number of parameters that can be simultaneously analyzed, restricting their utility. Spectral analysis technology promises a greater flexibility for panel design, a higher number of analyzed parameters, and more accurate visualization of results. Our standard all-in-one flow cytometry panel uses all possible fluorescent detectors of the flow analyzer BD Fortessa, which permits the differentiation of the main immune cell populations in the tumor, such as T cells (CD4+, CD8+, Treg), B and NK cells, macrophages (M1/M2), MDSCs (granulocytic and monocytic), and dendritic cells. However, in addition to immune cell frequency, it would also be interesting to investigate cell surface activation and memory markers like PD-1, CD40, CD44, or CD62L or intracellular activation markers like IFN-gamma, perforin, granzyme B, or Ki67. The 32-channel spectral flow analyzer SP6800 ZE from Sony allows the combination of both. Flow cytometry data of different tumor samples will be presented, as well as the comparison of the results and resolution of both flow analyzer systems. Auto-fluorescence of cells, which causes fluorescent signal contamination of other fluorescent markers in the conventional flow cytometer, can be subtracted using spectral technology to see the true fluorescent population, thereby allowing dim signal detection in rare populations. A comprehensive overview will be given regarding the pros and cons of the spectral analyzer system and difficulties switching from the one system to the other. Citation Format: Philipp Metzger, Muriel Malaisé, Cynthia Obodozie, Holger Weber. Benefit of using a spectral flow analyzer for the analysis of immune cell populations in tumors [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 629.