iLite® CD3 Effector Cell/ T-Cell Activation Bioassays

T-Cell Activation in vivo
A mechanism of cell-mediated immune defense

T cells play a central role in cell-mediated immunity and can mediate long-term, antigen-specific, effector, and memory responses. 

T cell activation requires activation of two signals in T cells: recognition of antigen through the TCR/CD3 complex and CD28 co-stimulation via CD80/CD86. Co-engagement of these receptors on the cell surface leads to intracellular signaling events and the activation of nuclear transcription factors such as Nuclear Factor of Activated T cells (NFAT), NF-kB, and AP-1.

To be activated, the T-cells must receive several signals through antigen-presenting cells and engagement of the CD3/TCR Complex. Additional signals are provided by co-stimulatory molecules and their interactions. Many bi-specific antibody-based therapeutics utilize this activation pathway to induce a targeted immune response from the body's own T-cells.  The iLite T-cell activation product line offers a convenient and powerful way of measuring the efficacy of antibodies to elicit this effect in vitro.  

Luminescence is based on the crosslinking of CD3 complex, antibody, and target

The T-cell activation process is triggered when the effector cell interacts with a drug antibody bound to its target cell. The CD3 complex on the surface of the effector cell binds to the CD3 part of the antibody, thus creating a bridge between the effector and target cells.

Following the formation of this bridge, the engineered effector cells will, instead of activating the T-cell as in the in vivo situation, produce luciferase through an intracellular pathway and generate luminescence exclusively from this cross-linking and signaling. The strength of the luminescence correlates to the ability of the drug to activate the T-cells. 

A normalization gene for easy comparisons between samples

Like most iLite cell lines, the iLite CD3 Effector Cells also have a secondary luciferase readout from a luciferase gene expressed under the control of a constitutive promoter. This enables normalization of each individual readout according to cell number and compensates for potential matrix effects.

Novel biologics formats, including bispecific antibodies and CAR-T cell therapies, require robust, reproducible measurement of T cell activation.

The iLite CD3 T-Cell activating  Reporter Bioassay is designed for evaluating CD3 mediated T-cell activation, exhibits greatly reduced variability, and offers an easier workflow than traditional T-cell activation Assays. 

 Applications such as

• Potency lot release and
• Antibody screening
• Assessing the comparability between innovators and biosimilars