Chemotaxis assay

Chemotaxis is responsible for shaping the tumor immune infiltration, and in particular for the control of the recruitment of effector T cells (CD8+), regulatory T cells (Tregs), Th17, Tumor associated macrophages (TAMs) or dendritic cells (DCs). Explicyte offers to evaluate the migration caused by a drug candidate with possible chemoattractant properties using  IncuCyte® live content imaging, which unlike conventional single time-point chemotaxis assays,  allows to monitor in real time the migration. Furthermore, the migration of rare cells can be studied as the behavior of few individual cells can be tracked. 

Chemotactic migration of T-cells by live-cell imaging. Chemotactic migration of T-cells toward the chemoattractant SDF1α, visualized in real time on an 96-well cell migration plate. Transmembrane migration of T-cells was imaged and quantified using the IncuCyte®  live cell analysis system. The kinetic read out reflects the loss of cells from the upper membrane surface.

Assay Principle

 

  • T cells (primary or Jurkat cell line) are seeded onto a transluscent perforated holder (96 well-format).
  • The bottom face of the holder is immersed in the migration medium.
  • The cells at the upper face of the holder are continuously imaged and quantified.
  • Migration/Chemotaxis is monitored as the cells progressively disappear from the upper face of the holder while they migrate through the perforated holes.

Representative Results

Chemotaxis SDF-1

Differential modulation of migartory properties of activated T cells towards two chemoattractants by a CXCR4 antagonist.  Measuring the loss of cell area on the top of the membrane shows that anti-CD3/CD28 activated T cells migration towards CXCL11 and CXCL12 (SDF-1α) is differentialy modulated by the selective CXCR4 antagonist, AMD3100, which inhibits chemotaxis toward CXCL12 (IC50 = 279 nM), with no effect on CXCL11-mediated chemotaxis.

Chemotaxis - Reproducibility data

Precision and reproducibility of the monitoring of the migration of Jurkat T cells towards CXCL12. Jurkat cells were plated at a density of 5,000 cells per well in the holder of the migration plate. Two-fold dilutions of SDF-1α (n=8 per concentration) were added to the lower compartments.

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Explicyte's in vitro services for the screening of cancer immunotherapies include a range of kinetic cell-based-assays for the assessment of cancer cell migration, also called real-time chemotaxis assays.