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Mixed Lymphocyte Reaction (MLR)

The mixed lymphocyte reaction assay (MLR) is a method to measure the immunomodulatory effects (inhibit an ongoing immune reaction ) or immunogeneticity (induce of amplify a immune reaction) of a drug, and is commonly used to test the efficacy of immunotherapeutics, vaccines and other immunomodulators. In the MLR assay, mononuclear cells from two genetically distinct individuals are co-cultured resulting in cell activation, DNA synthesis and proliferation. MLR occurs due to the incompatibility of the antigenic determinants on the surface of the two cell populations. Typically, antigen presenting cells like Dendritic cells (inducer cells) are isolated form one donor while T-cells (responder cells) are also isolated but from a genetically unrelated donor (One way MLR).  Clients request this assay to detect lack of immunogenicity of their product in a one-way MLR test sample cultured with donor lymphocytes where only one cell type proliferates. Alternatively, enhancing or inhibiting effects of a compound are tested in the two-way MLR co-cultured with two donor’s mononuclear cells where both donor cells are allowed to proliferate. MarinBio quantifies cytokines synthesized during an MLR to judge the various pathways as well as measuring cell proliferation. Alternative readouts are DNA synthesis, and cell-specific activation markers.


Example of PBMCs (Peripheral Blood Mononuclear Cells) incubated for 6 days.

Left: One PBMC donor only. Right: Two allogenic PBMC donors. Note the large aggregate of cells due to the stimulation of proliferate from immunologically mismatched peripheral blood mononuclear cells.

FLOW Analysis of MLR

PBMC from Donor A is stained with cell trace violet and cultured with or without PBMCs from Donor B. FLOW analysis shows the cell size with forward scatter. Left Panel: Donor A cells stained with Cell Trace Violet (CTV) show only 0.7 percent of the total population is proliferating. Right Panel: Donor A cells stained with CTV incubated with unstained Donor B cells, show 5.7 percent of the total Donor A cells are proliferating. The Stimulation Index (SI) for this MLR is 5.7/0.7 = 8.0.

Increasing Cell Size In Forward Cell Scatter

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The MLR assays offered by MarinBio enable clients to understand and monitor any significant immunomodulatory effects of client drug candidates, for example within the tumor microenvironment and/or in combination with other immune modulators, including checkpoint inhibitors. MarinBio offers clients customized programs and high-quality data which can satisfy the FDA and give clients high confidence in the selection of lead drug agents. MarinBio has expert in vitro immunologists with pre-clinical and clinical immuno-biology and immuno-oncology experience. Our MLR services and assays can be specifically customized to client requirements.


MarinBio scientists have performed hundreds of MLR experiments using donor blood obtained on site, as well as with monkey, rodent and other species. We track our human blood donors for repeat analyses and can use frozen pairs of cells with convenient and reproducible MLR results. We are your scientists. We work with you to solve your needs. We submit results of each experiment in real time with all methods, materials, raw and analyzed data for optimal progress on the project.