Quantifying the impact of a CD19-BiTE combining flow cytometry & real-time cell analysis

The ability to quantitatively correlate T cell biomarker expression/secretion with  target cell killing is critical in both basic and applied studies of tumor immunology. Coupling flow cytometry with multiplex bead-based immunoassays enables quantitative measurement of multiple secreted targets in a single sample simultaneously. In this Application Note, we demonstrate a workflow involving the use of an xCELLigence Real-Time Cell Analysis (RTCA) instrument in conjunction with a NovoCyte flow cytometer to study both CTL-mediated destruction of tumor cells and the corresponding secretion of cytokines and cytolytic proteins by CTLs. Representing a promising new class of therapeutics, bispecific T-cell engagers (BiTEs) harness the power of the adaptive immune response by enhancing the ability of CTLs to specifically recognize and eliminate tumors. For example, CD19-BiTEs  are designed to simultaneously bind CD3 on CTLs and CD19 on cells of B cell lineage, thereby enhancing the efficiency of CTL effector functions against various B cell derived tumors. Herein, the enhancement effects of CD19-BiTEs have been evaluated using three distinct assays. While target cell death was monitored using an RTCA impedance assay and a flow cytometry-based cytolysis assay, secretion of cytokines and cytolytic proteins was quantified in a bead-based multiplex flow cytometry assay.

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