Multiplex profiling of human and mouse cytokines in humanized mouse models 9297

Abstract Description Humanized mice enable the study of human immune responses, vaccine efficacy, and immune-related diseases. However, the cross-species homology of immune-related proteins complicates distinguishing between human and mouse homologues in the same sample. Reliable tools to differentiate these homologues are crucial for using humanized mouse models. We developed a multiplex assay with mouse- and human-specific antibodies against cytokines. Specificity was confirmed by testing mouse and human recombinant proteins. This kit was optimized for mouse samples, using mouse-analyte capture beads, while human-analyte capture beads showed no reaction. To measure human cytokines, we spiked human PBMC supernatant into mouse samples, detecting the exogenous human analytes specifically on the human beads. We validated this platform by analyzing biospecimen fluids from immunodeficient NSG mice bearing orthotopic human glioma or bladder cancer xenografts treated locoregionally with tumor-targeting or non-targeting human CAR T cells. In both models, peripheral circulating levels of human-derived cytokines were lower in cohorts treated with locoregional CAR T cell administration compared to those receiving systemic administration, highlighting the applicability of this multiplex immunoassay kit in two different visceral compartments. This study demonstrates the feasibility of measuring mouse and human proteins simultaneously, providing a tool for advancing research in humanized mouse models. Topic Categories Tumor Immunology: Checkpoints, Prevention, and Treatment (TIPT)