This article from Molecular Devices explores using the CellXpress.ai cell culture system for cell culture automation.
Sponsored Content by Molecular Devices UK LtdReviewed by Aimee MolineuxAug 5 2024 Discovery of efficient anti-cancer drugs and drug combinations is essential for therapy success. Therefore, it is crucial that new methods for efficient drug efficacy testing are developed to support the discovery of new viable therapeutic targets.
For the endpoint assay, the spheroids were stained with a mixture of the Hoechst nuclear stain and viability dyes Calcein AM and EtHD. The spheroids were then subjected to imaging and analysis for spheroid size and live-dead cell scoring. Moreover, the ATP content was measured using a CellTiter-Glo assay. Luminescent read-outs were acquired using the SpectraMax® iD3 Multi-Mode Microplate Reader.
Analysis in TL was completed using IN Carta® Image Analysis Software’s SINAP model and analysis in FL was run using CME analysis protocol. Compound addition: 48 hours after plating, 50 µl of media was taken out and 2x concentrations of compounds in the volume of 50 µl were introduced via the “different media” protocol.
To properly define spheroids sizes and intensities with various channels, CME analysis protocol in IN Carta® Image Analysis Software was applied. Observed shifts in spheroid area and average intensities as well as ratios of live/dead average intensities were applied to assess any compound effects. For a comprehensive phenotypic characterization of cells, it is advisable to use the ImageXpress® Micro Confocal system.
Spheroids were imaged using CellXpress with the DAPI, FITC, and Texas Red channels, with a 10x or 4x objective. Image analysis was carried out using CME . Spheroids were imaged using viable fluorescent channels DAPI, FITC and Texas Red, using best-focus projection images from Z-track of images captured at 10x magnification.
Figure 5. After compound treatment for 3 days, spheroid samples were tested for ATP content using CellTiterGlo reagent for 3D samples. EC50 for Staurosporin was 0.0 μM, for Paclitaxel 0.5 μM. Data for Doxorubicin were ambiguous due to the possible contribution of compound into Lumi signal. Image Credit: Molecular Devices UK Ltd
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