Topic 1:

Insight on Development & Validation of B Cell ELISpot Assay for Ag-specific B cells as Target Engagement (TE) in compliance with BMV Guidelines: Currently no standardized protocol for regulatory qualification and validation of the assay; Consideration on Critical Reagents & Parameters for validation; Importance lot-to-lot bridging for all critical reagents to ensure consistent performance across different batches; Continuous external validation programs are crucial for assessing inter-laboratory harmonization, B cell assay optimization, and the standardization.

Topic 2:

Lesson Learned from Validation of FluoroSpot Assays to assess Cellular Immune Responses: No formal Guidance on how to validate FluoroSpot assays; Considerations for validation parameters/procedures since validation parameters need to be assessed differently between clinical and non-clinical studies; Challenges with production of Positive Controls (PC) for multi-analyte assays: How to identify an appropriate PC? Considerations on identification of Positive Responses by different evaluation methods and application of  Distribution Free Resampling Method.

Topic 3:

Evaluation of the Complementarity ELISpot & Intracellular Cytokine Staining (ICS) and Concordance of Results based on specific assay design, validation, and execution: ICS differentiates from ELISpot with the ability to phenotype the cells producing cytokine to specific peptide pools informing the nature of immune response induced; ICS can multiplex efficiently, ELISpot can be multiplexed with DualSpot or FluoroSpot; ICS can assess polyfunctionality in cells where higher polyfunctionality indicates higher quality of the immune response induced by vaccines or therapeutic interventions.

Topic 4:

Overcoming one of the Biggest Challenges with Clinical Flow Cytometry: The Sample Very Short Lifespan; Majority of samples being unusable 48 hours after collection; Massive amounts of effort are expended to move samples from their collection site to an analysis facility; Current best model is to move samples to regional labs for analysis; Testing New Reagents that could be used bedside as an additive to preserve the whole blood clinical samples, allowing them to be frozen and batched shipped and analyzed.

Topic 5:

Challenges with uniqueness of the Sensitivity for Flow Cytometry Assays and its Utility in Data Interpretation: What types of flow cytometry assays need a sensitivity assessment? These include High Dimensional Flow Panels, Functional Flow Assays and Receptor Occupancy (RO) Assays. What measurement units should be used to define the sensitivity thresholds, frequency, absolute count or MFI? How to apply the assay limit/threshold to interpret different types of flow cytometry data sets?

Topic 6:

Challenges in Automating Flow Cytometry for Clinical Sample Testing: Developing an end-to-end process within a GxP framework; Automating clinical sample processing, instruments, available platforms, and custom-built systems; Automating  regent preparation and cocktailing and assessment of several different platforms; Automating Supervised Automated Pipeline, addressing requirements, and resolving encountered issues; How can Unsupervised Approaches be used to supplement classical data reporting form clinical studies?

Topic 7:

Challenges in Establishing Analytic Performance Fit-for-purpose Criteria in Flow Cytometry: Examining ways of determining, defining and documenting the purpose of testing for new and existing assays; Walking through selecting development and validation experiments using white papers recommendations on analytic validation for flow cytometry; Common scenarios for more/less stringent performance criteria than typically used in a fit-for-purpose fashion; Leveraging the iterative process to refine fit-for-purpose performance.

Topic 8:

New Progresses in Functional Biomarker Assays using Flow Cytometry: Providing evidence of drug PD impact through measurement of changes in biomarker expression, proof of drug MOA, enabling clinical development decisions; Informing clinical decisions through understanding  impact of drugs on cellular responses; Target Engagement (TE) Assays report drug binding and inhibition of targeted signaling pathways to inform dosing decisions; Measurement of TE & PD for understanding depth & duration of drug coverage.

Topic 9:

Novel Techniques to Support Therapeutic Development with Mass Cytometry: Current gaps in translating tissue specific targets from circulating cells and the use of flow cytometry to fill these gaps; Using Imaging Mass Cytometry to deep phenotype and determine Target and Biomarker information within tissues. Leveraging imaging mass cytometry to look at RNA and protein in the context of mRNA therapeutics. Future directions for imaging mass cytometry.

Topic 10:

Implementation of Mass Cytometry & Nanostring Qualified Assays to monitor Differential Expression levels of  immune or metabolic genes and changes in cell-associated proteins of patients with advanced solid tumors; Mass Cytometry revealed statistically significant changes in expression of PPAR-α associated proteins the RNA of which were not detected to change by Nanostring; These findings point out the value of interrogating patient materials with multiple qualified assays to identify potential PD Biomarkers and utilizing statistical methods to corroborate associations with drug exposures.

Topic 11:

Advanced Applications of Flow Cytometry and Imaging Cytometry to advance development of Novel Modalities: Characterizing cell-binding properties of Target-Specific Antibodies: Antibody Binding Capacity (ABC) and measurements of apparent affinity; Measuring LNP-RNA Membrane-binding and uptake in cell populations of interest to inform on cellular mechanisms driving efficient delivery; Building and characterizing PrimeFlow RNA Flow Cytometry Assay for cell-specific simultaneous analysis of LNP-mediated RNA delivery and transgene expression.

Topic 12:

Latest Technological Developments in High Dimensional (HD) Flow Cytometry: Spectral Cytometry, Mass Cytometry, High Parameter Flow Cytometry: How these platforms have enhanced the capacity for multi-parameter analysis: Address complexities of analyzing data, issues related to data visualization, dimensionality reduction, application of advanced computational; Exploring future directions, integration of single-cell genomics and proteomics, development of standardized data analysis pipelines, translation of assays into clinical diagnostics and personalized medicine.

Topic 13:

Pioneering the Development of a Minimal Residual B-cell (MRB) Assay based on sequencing of the IgH locus to enable Polyclonal B-cell Counting Inspired by the use of Molecular Assays to track Malignant Cell Populations in oncology; Comparison between Flow Cytometry & Molecular MRB approaches in the context of assay accuracy (relative to cytometry) and precision; Molecular is comparable to Cytometry but requires bioinformatic filters to remove aberrant IgH sequences and house-keeping genes to quantitate total number of nucleated cells counted.

Topic 14:

Latest Advances of the Modular Approach and how this approach can change the way Flow Cytometry is applied in Clinical Trials: Considerations and Pro/Cons on the recent evolution of Flow cytometry technology specifically in the clinical settings; Focusing on the advantages of High Dimensional Flow Cytometry Modular Assay; strategies to successfully handle the validation in collaboration with CRO; What's next for the development of the modular approach?

Topic 15:

Innovative Development of Cell-Based Assay (CBA) to Evaluate and Characterize the Impact of a PI3K-gamma Inhibitor on macrophage suppressive phenotype within the Tumor Microenvironment: Myeloid Cells are among the most prevalent cell types in inflamed and non-inflamed tumors and may play a role in resistance to immune checkpoint blockade; Results: Induced significant inhibition of tumor growth, polarized immunosuppressive M2 macrophage phenotype, increased phagocytosis measured by bead uptake in myeloid cells.

2025 White Paper on Cell-Based Assays - Latest Advances, Challenges and Solutions from Internationally Recognized Key Opinion Leaders

Consensus & Conclusions on Cell-Based Assays - Latest Advances, Challenges and Solutions from Internationally Recognized Key Opinion Leaders for 2025 White Paper