Mixed Lymphocyte Reaction (MLR)

Although sometimes mixed in their names, different sets of in vitro PBMC assays are commonly used to analyze innate or adaptive immune activation, as well as alloreactivity studies.:

• Peripheral blood mononuclear cell (PBMC) immunogenicity assays are commonly used to evaluate the ability of a test article (e.g., a biologic, cell therapy, or donor cells) to stimulate human immune responses in vitro.
• The mixed lymphocyte reaction (MLR) is a standard approach in which PBMCs from different donors are co-cultured to measure alloreactive T-cell activation driven by HLA mismatch.
  • In a one-way MLR, PBMCs from a responder donor are cultured with stimulator PBMCs that have been rendered non-proliferative (typically by irradiation or mitomycin C treatment), allowing only the responder cells to proliferate and produce cytokines. This format is useful for assessing the immunogenic potential of a specific donor cell population or a therapeutic product.
  • In a two-way MLR involves co-culturing untreated PBMCs from two donors, permitting bidirectional recognition and activation of both cell populations. Two-way MLRs model mutual alloreactivity and are often used to study general donor compatibility or overall immune responsiveness. Readouts for both assays typically include T-cell proliferation (e.g., CFSE, CTV or CPD dilution, thymidine incorporation), activation markers (CD25, CD69), and cytokine production (e.g., IFN-γ, IL-2).

Applications of MLR in drug discovery and immunology

• Assessing immunogenicity of new compounds
• Assessing immunogenicity of various cellular therapies
• Evaluating the efficacy of cancer immunotherapies
• Studying autoimmune diseases and inflammation
• Testing vaccines and other immunomodulatory drugs