Abstract

Microraft Array Platform for the Selection of Lymphocytes Based on Target-Cell Killing

Nancy Allbritton, Frank and Julie Jungers Dean of the College of Engineering and Professor of Bioengineering, University of Washington in Seattle

Adoptive cellular therapy (ACT) is an emerging therapeutic in which cytotoxic T lymphocytes (CTLs) that recognize tumor cell epitopes are introduced into patients providing immunity against the cancer cells. For ACT to succeed, CTLs with high tumor-killing efficiency must be identified, isolated, and characterized. Current technologies do not enable simultaneous assay of cell behavior or killing followed by recovery of the most efficient killer cells. A microraft array technology that measures the ability of individual T cells to lyse a population of target cells followed by sorting of living cells into a multi-well plate for expansion and characterization was developed.  The microraft array platform was combined with image processing and analysis algorithms to track and monitor killing assays over many hours. Automated cell collection was incorporated into the platform for facile cell collection from the array. As a proof of principle, human T cells directed against an influenza antigen were co-cultured with antigen presenting target cells on the microraft arrays. Target cell killing was measured by tracking the appearance of dead cells on each microraft over time. Microrafts with a single CTL demonstrating the greatest rate of target cell death were identified, cloned, and influenza-antigen reactivity confirmed. The platform is readily modified to measure the antigen-specific activity of individual cells within a bulk CTL culture or the cell heterogeneity within a population of gene-engineered T cells.