Immune cell activation assessment

Modulation of the immune system function aiming at enhancing the potential of immune cell responsiveness is one promising challenge for cancer therapy. During the immune response, T cells become activated and undergo clonal expansion followed by differentiation into effector cells and induction of cell-mediated cytotoxicity and/or cytokine release.
Understanding and modulating the mechanisms underlying activation, proliferation & clustering of immune cells is key for the development of innovative approaches to promote immune cell function and tackle tumor progression.
One of the most common ways to measure immune cell activation in vitro is the assessment of T cell proliferation upon antigen- or TCR-mediated stimulation. In this respect, Explicyte is offering valuable in vitro PBMC- and T cell-based 96-w plate assays to evaluate the effects of immunomodulatory compounds/antibodies alone and in combination with anti-CD3 antibody, on immune cell activation. By live cell time-lapse imaging, treatment effects are evaluated in a kinetic and phenotypic manner on the immune cell proliferation and clustering, as well as functionally on the release of key effector cytokines.

Assay principle










 
  • Our 96-w plate assay is performed on immune cells from various origins (PBMCs, purified populations, …)
  • Real time visualization & kinetic monitoring of immune cell activation (proliferation) and aggregation (cell-cell clustering) by live cell imaging.
  • Cell confluence is measured, using an appropriate platform-associated algorithm & image analysis module, as a surrogate of immune cell proliferation.
  • Phenotypic immune cell proliferation and clustering measurement can be multiplexed with additional functional measures i.e. effector cytokine release.

Illustrative data

Real-time monitoring of immune cell proliferation
Real-time live cell monitoring of immune cell proliferation (a). Automated image analysis & quantification of PBMC proliferation (yellow mask) overtime. αCD3 (5µg/ml) activates PBMC and induces their proliferation. Real-time live cell immune cell proliferation in the presence of various concentrations of αCD3 (b). Time- and dose-dependent effect of αCD3 in inducing hPBMC proliferation monitored over a ~120h period. 

 

Real-time live cell monitoring of immune cell proliferation (a). Automated image analysis ...

PBMC activation triggers key cytokine release
CD3-induced PBMC activation results in a time- and dose-dependent increase of key proinflammatory effector cytokines e.g. IFN𝞬 and TNF𝞪. hPBMC are cultured, for 24 and 72h, in the absence and presence of increasing concentrations of 𝞬CD3. Released amounts of IFN𝞬 and TNF𝞪 are then quantified in supernatants using specific HTRF technology-based detection kits. 

CD3-induced PBMC activation results in a time- and dose-dependent increase of key proinflammatory ...

PD1 blockade promotes PBMC proliferation & activation
Time-lapse live cell monitoring of PBMC proliferation (a). Kinetic monitoring of cell confluence (as a surrogate) highlights that PBMC rapidly proliferate under anti-CD3 activation, and that this effect is enhanced by PD1 blockade.

Anti-PD1 antibodies – Nivolumab & Pembrolizumab – promote αCD3-induced PBMC proliferation & activation (b). Treatment of αCD3-activated PBMC with increasing concentrations of each of anti-PD1 antibodies (72h) results in a dose-dependent IFN𝞬 release.  

Time-lapse live cell monitoring of PBMC proliferation (a). Kinetic monitoring of cell confluence ...