Immunotoxicity assays are laboratory tests used to evaluate whether a substance (e.g., drug, chemical, biologic, or nanomaterial) has adverse effects on the immune system. These assays assess how the immune system’s cells, functions, or signaling pathways are affected by exposure to the test compound.
Purpose: The primary purpose is to detect immunosuppression, which can increase the risk of infections or cancer, and immunostimulation, which may lead to autoimmunity, chronic inflammation, or cytokine storms. These assays also assess the potential for hypersensitivity and allergic reactions, as well as identify damage to immune organs such as the spleen, thymus, and bone marrow. Additionally, they are used to detect unintended immunomodulatory effects of biologics.
| Assay | Principle | Application | FDA Context |
|---|---|---|---|
| PBMC Cytotoxicity Assay | Measures the cytotoxic effects of test compounds on human PBMCs by quantifying markers of cell death, such as ATP depletion (indicating loss of viability) and LDH release (indicating membrane damage), in a high-throughput format | - Early in vitro screening of drug-induced cytotoxicity - Identification of off-target immune cell toxicity - Compound prioritization in preclinical development |
This assay is highly recommended for immunomodulatory or cytotoxic compounds to identify potential immune-related adverse effects to support early-stage safety profiling |
| PBMC Cytokine Release Assay | Measures inflammatory cytokines (e.g., IL-6, TNF-α, IFN-γ) released from PBMCs | Detects cytokine storm or immune overstimulation | Required for biologics (e.g., CAR-T, T-cell engagers, bispecifics); critical safety tool |
| T Cell Activation Assay | Flow cytometric analysis of CD69, CD25, IFN-γ expression | Characterizes immune activation or suppression | Supports immunotoxicity profiling under CBER guidance |