Abstract 3053: Comparative validation of neuroblastoma cell lines using flow cytometry and CyToF
Authors: Mulama David , Riveria Roman J. , McKinney Kimberly , Gandra Divya , Smith Kaitlyn , Tastet Nicholas , Sholler Giselle L. Saulnier 
Published: April 2023

Abstract
Introduction The validation of patient derived cell lines and other biologics has gained importance due to more frequent laboratory collaborations. Polychromatic flow cytometry has been the gold standard for detection and analysis of biochemical, structural complexity and characterization of cellular particles in biological systems. Despite the tremendous advancement in flow cytometry from the earlier simple, slow version of 1960s to more recent complex, fast spectral analyzers with comprehensive structural and functional immune profiling capability, this development has not matched the ever-increasing need for high throughput output to phenotype cells. Mass cytometry (CyToF) uniquely combines time-of-flight mass spectrometry with metal-labeling technology which can theoretically but simultaneously analyze 50+ parameters on a single cell. Despite the functional similarities in utility, mass cytometers and flow cytometers have different operating platforms that may impact their functional readouts . The goal of this study was to investigate and compare phenotypic readouts between flow cytometry and Cytof platforms using human neuroblastoma cell lines.
Methods To test this question, we utilized custom designed staining panels for both flow cytometry (fluorochromes-PerCp Cy 5.5, PE and Coralite 488 respectively) and Cytof (lanthanides: 155Gd, 151Eu, and 116Cd respectively) targeting CD56, Nestin and Synaptophysin, markers that are consistent with neuroblastoma using both five established neuroblastoma cell lines (BE2C, CHLA90, SMS-KCNR, SHSY5Y) and NGP) as well as four novel cells lines established in our laboratory derived from patient specimens (SL01277, SL01404, SL01255 and SL01287). We compared the percent expression of respective markers across cell lines between the two platforms.
Results Using both techniques we demonstrated that there was no difference in detection of viability in cells when validated under flow cytometry or CyToF (p&lt0.05). We also show extracellular staining of CD56 between the two platforms are comparable(p&lt0.05) across tested cell lines. Finally, we demonstrate that intracellular structures can be detected using both platforms with no significant difference (p&lt0.05).
Discussion and Conclusion The congruence and reproducibility in readouts between flow cytometry and CyToF analysis indicates that either assay can be used to study biological systems. Given the high dimensionality and versatility, then CyToF offers a robust platform that can be leveraged for immunophenotypic and functional studies of cellular material. Furthermore, CyToF due to its wide breadth of multiparametric measurements available will be an instrument of choice where the study of multiple parameters is desired.
Citation Format: David Mulama, Roman J. Riveria, Kimberly McKinney, Divya Gandra, Kaitlyn Smith, Nicholas Tastet, Giselle L. Saulnier Sholler. Comparative validation of neuroblastoma cell lines using flow cytometry and CyToF [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3053.