Arrays of single cancer cells were obtained by seeding renal cancer cells (RCC-26) on small adhesive micropads in an ibidi µ-Slide VI 0.4 (pattern size: 40 µm, pattern distance: 120 µm). After cell adhesion, JB4 T cells were applied in suspension and live cell imaging was performed for 30 hours, nicely illustrating the cancer cell killing by the specific T cells.
Data analysis was performed using FastTrack AI by MetaVi Labs. The array information helps to keep the cell number and positions constant thereby facilitating the analysis. The AI-supported algorithm determines over time whether a cancer cell is attached to a single adhesive pad or not. Like this, the T cell killing efficiency can be calculated.
Autologous T cells that express engineered antigen receptors (CAR-T cells) represent a promising new cancer therapy tool. The evaluation of quality, specificity, and killing efficiency (potency) of CAR-T cell populations is crucial for the development of potent and safe patient-specific CAR-T cell therapies.
In contrast to classical T cell potency assays (e.g., Chromium-51), live cell imaging allows to analyze T cell/cancer cell interaction in real time with single cell resolution.
However, analysis of confluent cell layers is very time-consuming and therefore not possible in high throughput screens. Additionally, variations in cell size and density require fluorescent labeling for automated T cell and cancer cell registration, which might alter the cell behavior.
To facilitate high throughput label-free analysis of T cell potency in a live cell imaging setup, we generated arrays of homogenously distributed single cancer cells. By combining optical analysis and advanced image processing, cytotoxic T cell activity over time on a single cell level can be evaluated without the use of any labeling.
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29 авг 2024
