Assessment of antigen-specific T cell recall responses in non-human primates using a composite AIM (cAIM) assay

Jan Marten Schmidt^1*Mayuri Prasad²Kenna Degner²Rebecca Schiavo²Allison Repic²Lauren Little²Laura Israel³Feng Chen⁴Regine Hansen¹Anette Christine Karle³Kees Leenhouts⁵Poul Sørensen^5,6Brian Mcintosh²Dominique Brees¹José M Carballido^1*

• ¹Novartis Biomedical Research, Preclinical Safety, Basel, Switzerland
• ²LabCorp Early Development Laboratories, Madison, United States
• ³Novartis Biomedical Research, Biologics Research Center, Basel, Switzerland
• ⁴Novartis Biomedical Research, DAx, Cambridge, MA, United States
• ⁵Allero Therapeutics, Rotterdam, Netherlands
• ⁶Department of Biomedicine, Aarhus University, Aarhus, Denmark

Characterizing antigen-specific T cell responses is essential for understanding the immunogenicity of biotherapeutics and mitigating drug-specific immune reactions. This study describes a flow cytometry composite Activation-Induced Marker (cAIM) assay for cynomolgus monkeys, that allows quantification of T cell recall responses to multiple antigens using up to ten AIM pairs. The procedure incorporates two composite metrics (cAIM-index and cAIM-score) that facilitates the summation of T cell recall responses into interpretable numeric values, reducing reliance on multiple graphical comparisons. The assay is compatible with human and mouse samples and can utilize peripheral blood mononuclear cells or whole blood. Additionally, the method is well suited for the mass cytometry platform, enabling the detection of antigen-specific CD4+ and CD8+ T cell recall responses while providing deep immunophenotype information and consuming minimal blood sample volumes. These protocols shall support preclinical and clinical immunogenicity assessments, advancing biotherapeutic development.