Journal: Communications Biology

Article Title: A statistical framework for high-content phenotypic profiling using cellular feature distributions

doi: 10.1038/s42003-022-04343-3

Figure Lengend Snippet: a A set of 65 compounds with diverse chemical structures and a wide range of ChEMBL target classes were selected to examine the performance of the marker panels. Hierarchical clustering using the Tanimoto distance metric shows pairwise chemical structure similarity. b The 384-well plate layout for chemical screening: control wells (DMSO treatment) are placed in a diagonal pattern, and a dilution series of compounds arranged horizontally with high to low concentration (left to right). The outermost rows and columns are excluded from scanning and analysis to mitigate edge effects due to evaporation. c Dilution series of 65 compounds and control wells were spread over two plates, with three replicates for each condition. The heatmap shows well-level cell counts in different plates. d Scatter plot showing cell counts in control (left panel) and treatment (right panel) wells, ordered by plate and replicate number (grey dashed lines correspond to min, max and median of control cell count). e Hoechst 33342 (nuclear DNA) stain distribution showing density curves for individual control wells (red) and aggregated control wells (global control, black, sample size: 265,638 cells). f Distribution of total nucleus intensity of cells after the treatment with increasing doses of Mitoxantrone (sample sizes from low to high dosage (n 0.078125 – n 5 ): 711, 669, 559, 372, 325, 363, 363 cells). g Cells treated with different compounds and concentrations (A = irinotecan 5 μM, B = monensin 0.3125 μM, C = rapamycin 10 μM, D = vincristine 10 μM) show a diverse distribution of total nucleus intensity (sample size: Control = 265,638, n A = 1544, n B = 1817, n C = 1849, n D = 1173 cells).

Article Snippet: The source plates containing serial dilutions of the compound and DMSO controls were prepared by combining McCoy’s 5A medium (no FBS added) and various concentrations of test compounds in 384-well plates (Corning, Cat. number: CLS3657), with a specific diagonal pattern of controls (DMSO only).

Techniques: Marker, Concentration Assay, Evaporation, Cell Counting, Staining

Journal: Assay and Drug Development Technologies

Article Title: High Content Screening Characterization of Head and Neck Squamous Cell Carcinoma Multicellular Tumor Spheroid Cultures Generated in 384-Well Ultra-Low Attachment Plates to Screen for Better Cancer Drug Leads

doi: 10.1089/adt.2018.896

Figure Lengend Snippet: Characterization of HNSCC MCTS morphology and viability in Ultra-low Attachment Plates. Eleven HNSCC cell lines and Het-1A cells were seeded at 2,500 cells per well in 384-well ULA-plates and cultured for 3 days. MCTSs were then stained with the live cell CAM and dead cell EHD reagents, and 4 × images were acquired on the IXM in the TL, FITC, and Texas Red channels. Grayscale TL images are presented along with color composite fluorescent images of live cell CAM and dead cell EHD staining depicted as green and red, respectively. Representative images from multiple independent experiments are presented. All scale bars represent 300 μm. CAM, Calcein AM; EHD, Ethidium Homodimer; HNSCC, head and neck squamous cell carcinoma; IXM, ImageXpress Micro; MCTS, multicellular tumor spheroid; TL, transmitted light; ULA-plates, ultra-low attachment microtiter plates.

Article Snippet: Generation of HNSCC MCTSs in Ultra-Low Attachment Microtiter Plates We have previously described the generation of MCTSs after seeding several HNSCC cell lines into 384-well U-bottomed ultra-low attachment microtiter plates (ULA-plates; Cat. No. 4516; Corning, Tewksbury, MA).

Techniques: Cell Culture, Staining