46 Antigen density quantification by mass cytometry using quantum simply cellular bead-based calibration
Year: 2026
Type: conference-abstract
Abstract: Background Quantifying cell surface antigen density is critical for the development of biologics, antibody-drug conjugates (ADCs), and other targeted therapies. For ADCs, antigen density influences target engagement, internalization, payload delivery, and therapeutic efficacy. For T cell engagers and CAR-T cell therapies, antigen abundance can affect immune activation and treatment potency. Equally important is measuring low-level antigen expression on healthy tissues, where unintended target engagement may contribute to off-tumor toxicity. Despite its importance, quantitative antigen density analysis has remained technically challenging and inaccessible for many drug developers, limiting its use in target selection, patient stratification, and translational research.Methods We developed an antigen density quantification workflow for mass cytometry using Quantum Simply Cellular (QSC) beads with known antibody binding capacities (ABC). QSC beads were stained with the same metal-conjugated antibodies used to profile experimental samples, including peripheral blood mononuclear cells (PBMCs) and whole blood. Beads were acquired by mass cytometry to generate marker-specific calibration curves relating instrument signal to antibody binding capacity. These calibration models were then used to convert cellular mass cytometry signal into absolute antigen density, reported as molecules per cell.Results QSC beads stained with an anti-CD19 metal-conjugated antibody generated a robust calibration curve relating mass cytometry signal to antibody binding capacity. Linear regression of the calibration curve demonstrated excellent performance, with an R 2 > 0.98 (figure 1). This calibration model was subsequently used to quantify CD19 antigen density on cells by converting mass cytometry signal into standardized estimates of molecules per cell. The workflow integrates seamlessly with routine mass cytometry sample preparation and acquisition, enabling quantitative comparison of antigen density across experimental samples. Evaluation of additional markers is currently underway to expand the applicability of this approach across a broader range of therapeutic targets.Conclusions We present a straightforward, bead-based workflow that extends the utility of mass cytometry beyond relative expression profiling by enabling quantitative measurement of cell surface antigen density. By converting instrument signal into standardized estimates of molecules per cell, this approach provides drug developers with quantitative measurements of target abundance to support target selection, therapeutic development, biomarker discovery, and translational research.Ethics Approval This study was reviewed and approved by WCG IRB (# 20250604). All participants provided written informed consent prior to enrollment, in accordance with the Declaration of Helsinki.Abstract 46 Figure 1CD19 antigen density
Language: English
Subfield: Immunology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,490
OpenAlex ID: W7225309654