Cytotoxicity Assays
Cytotoxicity assays are foundational to in vitro toxicology services and play a critical role in the early-stage evaluation of drug candidates, chemicals, cosmetics, biologics, and medical devices. They act as the First Line of Safety Assessment.
Purpose: Detect if a compound is toxic to cells by measuring cell viability, membrane integrity, or metabolic activity.
| Assay | Principle | Application | FDA Context |
|---|---|---|---|
| MTT / MTS Assay | Mitochondrial dehydrogenase activity (colorimetric) | General cell viability | Widely accepted for early screening |
| LDH Release Assay | Quantifies membrane damage via lactate dehydrogenase leakage | Detects necrosis/cell lysis | Commonly used in IND-enabling studies |
| ATP Assay (e.g., CellTiter-Glo) | Measures intracellular ATP as viability marker (luminescence) | High-throughput, sensitive | Recommended for biologics with immune or cytotoxic targets |
| Trypan Blue Exclusion | Dye exclusion by live cells | Manual viability check | Traditional assay, still used in some IND packages |
| Annexin V / PI Flow Cytometry | Detects early/late apoptosis and necrosis | Mechanism-based cytotoxicity | Recommended for biologics and cell therapy products |
| Caspase-3/7 Activity Assay | Measures executioner caspase activation in apoptosis | Apoptosis pathway specificity | Common in cell therapy preclinical assessment |
| Real-Time Cytotoxicity Assays | Continuous impedance or imaging-based monitoring (e.g., xCELLigence, Incucyte) | Dynamic response profiling | Informative in biologics and immunotherapies |
| Live/Dead Cell Staining | Calcein AM + Ethidium homodimer or similar dyes | Fluorescence imaging of viability | Often used in cell therapy, 3D models, and co-culture |