"Lethal kiss” on a dielectrophoresis (DEP) based Lab-on-a-chip device: two-colour real-time determination of cytotoxic T-lymphocyte activity.

Cytotoxic T-lymphocytes (CTL) are of great importance for the control of the immune response against tumor onset and progression. It is well known indeed that CTL of cancer patients can exhibit the ability to lyse tumor cells. This feature is of great relevance for the development of immunotherapy of tumors. Accordingly, the analysis of the cytolytic activity of isolated CTL populations displaying cytotoxic activity against tumor cells is of great clinical impact, since the isolation of highly cytoxic CTL clones is a complex and long procedure needed of step by step validation of the enrichment achieved. In this respect, dielectrophoresis (DEP) based Lab-on-a-chip technology could represent a very appealing approach, since it allows the manipulating of large numbers of single cells or cell populations. Here we present two dielectrophoresis-based Lab-on-a-chip platforms useful for the analysis of CTL-mediated cell lysis. One platform (the SmartSlide) displays parallel electrodes and generated cylinder-shaped cages within which CTL and target cells are entrapped. The second platform (the DEParray) generates spherical DEP cages that can entrap clusters composed of several CTLs and a single (if present) target cell. We present a two-color staining approach to label target cells to the aim of a real-time determination of the cytotoxic activity of T-lymphocyte precursors in these two (DEP)-based Lab-on-a-chip platforms. We demonstrate that these devices can be used to manipulate CTL clustered to target cells with the aim to develop an efficient methodology to quantify CTL-mediated cytolysis and identification of CTL clusters or clones exhibiting high cytotoxic activity. The proposed approach is not radioactive, fast and is suitable with the use of low numbers of target cells.