In Vitro Immunogenicity Assessment
The key process in T cell dependent antibody production is peptide presentation by antigen presenting cells to T cells, in combination with co-stimulatory signals and cytokine secretion. These three signals will stimulate the T cells to proliferate and differentiate, in order to be able to help the B cells to produce antibodies towards the antigen. Antibodies produced towards the administered drug (anti-drug antibodies or ADAs) are the main clinical immunogenicity problem. These ADAs can lead to a decreased or enhanced clearance of the drug, or can have no effect at all. If the drug mimics an endogenous counterpart, the ADAs might also bind the endogenous protein, which can lead to severe consequences for the patients’ health. The process from antigen presentation until ADA production cannot (yet) be tested in vitro. Therefore, T cell proliferation is can be used as a surrogate marker to measure the immunogenic potential of a drug candidate in development.
T cell activation assays have a long tradition in vaccine testing (wanted immunogenicity), but are also employed to asses possible unwanted immunogenicity induced by therapeutic proteins.
ImmunXperts offers several types of T cell activation assays:
- Innate immune response (Early reponse)
- Whole Blood Cytokine Release Assay: CRA are risk assessment tools for biotherapeutics possibly eliciting adverse pro-inflammatory clinical infusion reactions. Here, a small volume of anti-coagulated blood is incubated in the presence of the test product. After an optimised culture period, the plasma is collected and the secretion of cytokines can be assessed with ELISA or multiplex bead-based immunoassays.
- PBMC activation Cytokine Release Assay: In a different CRA format, cryo-preserved PBMC can be used instead of whole blood and cytokine secretion in the cell culture supernatant will be measured.
- Adaptive immune response (Late reponse)
- Whole PBMC assay: In the PBMC assay, complete test products are added to cultured peripheral blood mononuclear cells (PBMC). These cells contain all essential components to mount an immune reaction. After a period of incubation T cell activation is measured.
- DC-T assay: When test products interfere with the formation of an immune response, the DC-T assay type is preferred. Here, the test product is first loaded onto the surface MHC molecules of professional Antigen Presenting Cells (APC) and leftover product will be washed away. The APC will present all epitopes to autologous T cells and T cell activation is measured.
ImmunXperts has experience with several readout formats:
- Flow Cytometry: Multiparameter analysis on a single cell level
- Cytokine Profile: Capture beads allow the multiplexed analysis of more than 10 inflammatory cytokines
- ELISpot: Analysis of secreted molecules on a single cell level
For questions or to request a quotation, please contact us.