As the basic mechanism that allows our bodies to identify, target, and kill infected or damaged cells, cell-mediated cytotoxicity is an essential function of our immune system and one that scientists continue to explore to develop more advanced immunotherapies. 

By better understanding the role of T cells and natural killer cells as effector cells and different measurement methods, researchers can more accurately study the functions of these important immune cells in vitro. 

An 8-Minute Read

Take a few minutes to read this white paper for an overview of the types of cell-mediated cytotoxicity and various measurement methods. You will learn about:

  • Antigen-Specific Cell-Mediated Cytotoxicity
  • Antibody-Dependent Cellular Cytotoxicity
  • Natural Killer Cell-Mediated Cytotoxicity
  • Chromium-51 (51Cr) Release
  • Lactate Dehydrogenase (LDH) Release
  • Calcein Release
  • Staining to Detect Degranulation
  • Flow Cytometry


Measuring Cell-Mediated Cytotoxicity

1. Antigen-Specific T Cell-Mediated Cytotoxicity

Antigen-specific T cell receptors bind to MHC class I molecules; CD8+ T cells release perforin and granzymes 

Employed by cancer immunotherapies targeting immune checkpoints

2. Antibody-Dependent Cellular Cytotoxicity

Fc receptors on effector cells bind to antibodies on the surface of target cells; effector cells release cytotoxic granzymes

Utilized by certain antibody-based drugs, such as Herceptin®  and Rituxan® 

3. Natural Killer Cell-Mediated Cytotoxicity 

Requires neither antibody nor antigen expression; NK cells directly recognize target cells and release cytokines

Chimeric antigen receptors (CARs) engineered into NK cells as alternative


Measurement Methods

1. Chromium-51 Release

Target cells are labeled with 51Cr and release 51Cr through cell lysis. 

  • Sensitive — the release of 51Cr is easy to detect
  • Measures death of target cells, not death of killer cells
  • Some leakage of the label, higher background
  • Requires hazardous materials

2. Lactate Dehydrogenase (LDH) Release

Measures amount of soluble cytosolic enzyme released during cell death using colorimetric readout.

  • LDH is more stable than other enzymes
  • No label required
  • Release of LDH is not limited to the target cells
  • High background


3. Calcein Release

Target cells are labeled with highly fluorescent Calcein AM. Damaged and killed cells release Calcein into culture. 

  • Can be detected using fluorescent plate reader
  • Label is specific to target cells
  • High background

4. Staining to Detect Degranulation 

Directly labeled anti-CD107a added to T cells or NK cells prior to exposure to target cells. Allows for detection of antigen during degranulation. 

  • Only requires one label
  • Low background
  • May miss degranulation
  • Flow cytometer needed

5. Flow Cytometry

Target cells are labeled with a fluorescent viability dye to differentiate them from the effector cells. 

  • Deeper analysis
  • High sensitivity
  • Not as easily read in larger assays

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