CYTO 2026 Tutorials: Minimum standards and best practices to ensure reproducibility in longitudinal flow cytometry studies

Minimum standards and best practices to ensure reproducibility in longitudinal flow cytometry studies

Presenters:
Kathryn Hally, PhD, Senior Lecturer, University of Otago Wellington (Ōtākou Whakaihu Waka ki Pōneke)
Ana Longhini, PhD, Global Scientific Affairs Senior Manager, Sony Biotechnology Inc
Laura Ferrer Font, PhD, Scientific Solutions Manager, BD Biosciences
Megan McCausland, BSc, Scientific Advisor, Flow Cytometry, IQVIA Laboratories
Sam Small, Other, Senior Specialist, Flow Cytometry, Malaghan Institute of Medical Research

Abstract:
Highly reproducible flow cytometry assays are essential for generating robust data in longitudinal studies. These studies offer powerful insights into disease progression, treatment response, and immune dynamics. Their inherent complexity, however, demands meticulous planning and execution to ensure consistency across timepoints, instruments, operators and sites. Other critical factors - including accounting for batch effects, reagent performance, and sample handling and storage - also require tight control to reduce their impact on the integrity of cells or markers of interest. Further, as cytometry technologies continue to evolve, researchers are designing higher dimensional panels, integrating data from multiple instruments and sites, and using sophisticated analytical approaches; while promising, these advancements introduce new challenges for reproducibility. Notably, longitudinal study design is not one-size-fits-all; these must be tailored to the users’ expertise, resources, and the assay’s intended use.

To better address the challenges of executing longitudinal flow cytometry studies, we conducted two community surveys and organized two workshops at CYTO2024 and CYTO2025. Our objective was to gather diverse perspectives from the cytometry community and collaboratively define, in alignment with published guidance, a set of minimum requirements and best practices to support reproducibility in longitudinal studies. We explored critical elements such as assay validation, antibody and reagent lot control, ensuring consistent instrument performance, the necessity of an assay-specific quality control, and considerations for data analysis. In this tutorial, we will present this framework and highlight how adherence can significantly enhance assay reliability and interpretability in longitudinal flow cytometry studies.

Learning objectives:
Identify the key challenges involved in planning and executing a longitudinal flow cytometry study within the attendee’s own research context.
Detect common sources of variability within these studies, and select strategies to minimise their impact.
Apply the community-informed minimum requirements for conducting these studies, informed by practical examples of their implementation.
Develop and support complex longitudinal study designs involving multiple instruments, operators, and sites.

Keywords: Instrument Standardization, Experimental Design & Controls, Best Practices, Longitudinal Studies

CMLE Credit: 1.5