Application Note: Utilizing Mixed Lymphocyte Reaction (MLR) to Evaluate Checkpoint Inhibitor Therapies by Advanced Flow Cytometry

In the field of immunotherapy checkpoint inhibitors have been developed, which block the immune checkpoints that normally ‘switch off’ T cells. Future immune checkpoint targets are being explored particularly within drug discovery fields.

Immune checkpoints are signaling molecules that control the synapse between T cells and antigen presenting cells. Checkpoint inhibitor therapies target and inhibit the signals that ‘switch off’ T cells, leading to an up-regulation of T cell activation and tumor cell killing. The mixed lymphocyte reaction (MLR) mimics the T cell and antigen presenting cells synapse, by co-culturing immune cells from two individuals. MLR is used as a model to explore the effects of checkpoint inhibitor drugs in vitro.

Conventional techniques for analysis of MLR assays, such as ELISA and traditional flow cytometry and are often limited, as they often require the use of separate techniques and assays, for the measurement of proliferation and activation marker expression and for the measurement of cytokines. Hence manual collation of data from different techniques is required. In addition, large sample volumes at protracted sample throughputs is observed.

To overcome these difficulties, Sartorius has developed a simple, high-throughput workflow, where a validated suite of reagent kits is used in conjunction with the iQue® Advanced Flow Cytometry Platform, which includes integrated Forecyt® software, to provide a simple and flexible road map, for quantifying T cell response in an MLR assay. This has relevance to potential applications in immunotherapy research and drug discovery.

In this workflow, data is acquired at a high throughput rate and small sample volumes are used, to measure cell markers and cytokines in multiplex. Ranking of checkpoint inhibitor drugs, based on their activity in inducing T cell response is possible.

Hence libraries of potential checkpoint inhibitor therapeutics may be profiled for their activity, in minimal time.

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