T Cell Functional Assays

T Lymphocytes & T Cell Activation

T cells are central to adaptive, cell-mediated immunity. Originating in the bone marrow and maturing in the thymus, they differentiate into helper, regulatory, cytotoxic, or memory subsets. Upon encountering specific antigens, T cells are activated through a multi-signal process involving antigen recognition, costimulatory signaling, and cytokine release. Activated T cells regulate immune responses, support B cell activity, and directly eliminate infected or malignant cells.

At Axela, we provide a wide array of functional T cell assays to support immunology research, oncology, and drug development. Our expertise spans antigen-specific responses, checkpoint modulation, cytokine profiling, and advanced cytotoxicity models.

Our Services Include:

💠Activation & Proliferation Assays

    • Mixed lymphocyte reaction (MLR)

    • Anti-CD3/CD28 stimulation, PMA/Ionomycin, SEB, checkpoint blockade (anti-PD1, anti-CTLA4)

    • High-throughput flow cytometry and readouts (CFSE, BrdU, DELFIA)

💠Cytotoxicity & Response Assays

    • T cell-dependent cellular cytotoxicity (TDCC), including bispecific T cell engager (BiTE) assays

    • Tumor killing assays (TKA)

    • γδ T cell cytotoxicity assays

    • 3D spheroid T cell-mediated cytotoxicity

💠Checkpoint & Exhaustion Studies

    • Immune checkpoint blockade assays

    • T cell exhaustion and reversal assays

    • T cell differentiation and suppression assays (Treg/MDSC functional profiling)

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💠Cytokine & Phenotypic Profiling

    • Cytokine secretion (ELISA, Luminex, AlphaLISA, Lumit, ELISpot)

    • IL-2 and signaling assays (phospho-Flow, pSTAT5, etc.)

    • T cell surface marker expression (flow cytometry)

    • Chemotaxis and migration assays

💠Specialized Assays

    • On/Off target testing for TCR and cellular therapeutics

    • TAA-specific T cell response assays

    • Immune synapse formation assays

T cell activation requires three distinct signals provided by antigen-presenting cells (APCs).

  • Signal 1: Antigen recognition via the T cell receptor (TCR) binding to antigen–MHC complexes.

  • Signal 2: Costimulatory signaling through molecules such as CD28 and CD80/CD86.

  • Signal 3: Cytokine release from APCs, which drives T cell proliferation, differentiation, and effector functions.

Mixed Lymphocyte Reaction (MLR) Assay

MLR assay evaluates compounds that influence the interaction between antigen-presenting cells (APCs) and T cells, enabling assessment of T cell activation, deactivation, or reprogramming. It provides key insights into the efficacy of immunomodulatory agents and drug safety in immunology, oncology, and autoimmunity.

Axela’s MLR platform supports rapid identification of T cell–modulating agents, suitable for both biologics and small molecules. Using ultrassensitive immunoassays, multiplex cytokine profiling, and advanced flow cytometry, we deliver high-quality analysis across multiple endpoints, including proliferation, cytokine release (e.g., IL-2, IFN-γ), and cell-surface marker expression.

 

  • One-way MLR: Responder lymphocytes (e.g., CD4+ T cells) from one donor are stimulated by mitomycin C–treated stimulator cells (e.g., DCs) from another donor.

  • Two-way MLR: Lymphocytes from both donors stimulate each other, modeling broader allogeneic responses.

Our Services Include:

  • High-throughput format: 96–384 well plates

  • Robust, reproducible assays suitable for small to large-scale studies

  • Flexible design for single-agent or combination testing

  • Access to multiple allogeneic donors (PBMCs, CD3+/CD4+ T cells, monocyte-derived DCs)

  • Multiparametric readouts: cytokines, proliferation, and surface markers

  • State-of-the-art platforms: Luminex-200, FlexMAP 3D, CytoFLEX S flow cytometer (13-color, 4-laser)

  • Fast turnaround: quantitative results in as little as 4 weeks

  • Fully validated and quality-controlled with triplicate replicates

  • Comprehensive controls: isotype IgG, inhibitors, and stimulators

  • Over 20 years of hands-on expertise with expert analysis and reporting

Example Data

T Cell Activity Assay Example Data:
Representative data showing the effect of PD-1 blockade on T cell proliferation and cytokine production in a mixed lymphocyte reaction (MLR). Human CD4⁺ T cells were cocultured with allogeneic monocyte-derived dendritic cells (DCs) in the presence of either test antibody or isotype control.

  • Figure 1: BrdU incorporation and IFN-γ secretion demonstrate that anti-PD-1 treatment enhances both T cell proliferation and cytokine production compared to controls.

  • Figure 2: Flow cytometry analysis confirms increased proliferation of CD4⁺ T cells when treated with anti-PD-1 antibody.

These results highlight how MLR assays can be used to assess immunomodulatory activity and quantify functional enhancement of T cell responses.