Improved killing assay based on screening of Individual CAR cells

App Notes



CAR cell development stands out as a highly promising avenue in cancer research. Following the engineering of CAR cells, scientists face the crucial task of assessing the viability of these engineered cells. Traditional methods for viability assessment are often time-consuming and may overlook effective cells. In this Application Note, we showcase how LiveDrop instruments offer a rapid and straightforward solution for scientists to visually evaluate the efficacy of the CAR cell-based therapies they are developing.


Assessing the capacity of individual immune cells to eradicate cancer cells is crucial for predicting the effectiveness of personalized cell-based therapies. Immune cells eliminate both infected and malignant cells through their cytolytic activity. The dynamics of interaction between effector immune cells and target cancer cells exhibit heterogeneity in terms of the nature, intensity, and duration of their contact. Currently, the evaluation of Chimeric Antigen Receptor (CAR) cell efficacy against target cells primarily relies on co-culture assays. However, these assays provide only averaged end-point measurements, utilize extensive cell pools, and do not assess the cytolytic effect of individual cells or distinct cell subsets on target cells. Additionally, they fail to determine the proportion of active killer cells or the quantity of target cells eradicated by individual immune cells.

LiveDrop Innovation

By encapsulating a CAR cell with a target cell in a droplet (Figure 1), Modaflow enables the direct visualization of interactions between individual CAR cells and their targets. This feature, coupled with sorting capabilities, facilitates the assessment of efficacy with unprecedented clarity.

Cytotoxicity assays conducted within microfluidic droplets involve co-encapsulating cells and assay reagents into picoliter-sized droplets using the ModaFlow instrument. The EthD-III probe, utilized for cell death detection, will selectively color dead cells in red.
Figure 2: Macrophage cell-mediated cytotoxicity.(A) Cancer target cells labeled with CFSE dye (green).(B) Dead cancer cells (red).(C) Overlay image displaying dead target cancer cells in droplets.(D) Images showing various macrophage-to-cancer cell ratios in droplets.(E) Interaction between macrophage cell and target cancer cell. The droplet containing overlapping green and red fluorescence indicates the destruction of cancer cells. The red arrow indicates killer macrophage, while the green arrow indicates non-reactive macrophage.(F) Bar diagram illustrating the percentage of killer macrophage cells in 248 droplets.


As seen in Figure 2, encapsulating a CAR celland one or multiple cancer cells in a single droplet allows to visualize theelimination of the cancerous cells, if the CAR cell is effective. After 5H ofincubation, around 10% of macrophage cells induced cytotoxicity.


Screening and sorting individual CAR cells with ModaFlow provides visual evidence of their efficacy against targets, enabling rapid validation of engineered CAR cell functionality for scientists.

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