cfb  (Proteintech)

Journal: Journal of Molecular and Cellular Cardiology Plus

Article Title: Pericellular oxygen dynamics in human cardiac fibroblasts and iPSC-cardiomyocytes in high-throughput plates: insights from experiments and modeling ☆

doi: 10.1016/j.jmccpl.2025.100464

Figure Lengend Snippet: Overview of experiments and modeling of pericellular oxygen in HT 96-well plates. A. The experimental system and the experimental conditions are shown. In-incubator-operated optical imaging system included a camera, a lens, LED ring, and bellows extension to accommodate a glass-bottom 96-well plate. The ratiometric optical readout was calibrated in values of pericellular oxygen concentration over time. The experimental conditions included two cell types - adult cardiac fibroblasts, cFBs, and human iPSC-CMs, three cell densities, and four different culture medium volumes. B. Confirmation of cell growth on the optical oxygen sensors was done using a two-photon upright microscope with a water-immersion lens. Images of hiPSC-CMs on top of the sensor are shown at two different magnifications (25× and 40×). Color encodes depth in the lower magnification image, in which cells are labeled with Hoechst for nuclei and with ChR2-eYFP to highlight the membrane. The higher magnification image shows cytoskeleton (α-actinin) and nuclei (Hoechst); scale bar for the zoomed in version is 50 μm. C. The computational modeling framework is a combination of 3D physics-based simulations of reaction-diffusion in the wells of a HT plate and optimization algorithms to derive empirically-based models of pericellular dynamics in human cFBs and iPSC-CMs, grown in glass-bottom 96-well plates, for a variety of cases. Simulations and data processing were done in Matlab, LiveLink and Comsol.

Article Snippet: Human adult ventricular cardiac fibroblasts, cFBs (Cell Applications, Inc. 307V-75a) were thawed into a T75 flask per the manufacturer's instructions.

Techniques: Optical Imaging, Concentration Assay, Microscopy, Labeling, Membrane, Diffusion-based Assay

Journal: Journal of Molecular and Cellular Cardiology Plus

Article Title: Pericellular oxygen dynamics in human cardiac fibroblasts and iPSC-cardiomyocytes in high-throughput plates: insights from experiments and modeling ☆

doi: 10.1016/j.jmccpl.2025.100464

Figure Lengend Snippet: Experimental tracking of pericellular oxygen in human cardiac fibroblasts (cFB) over 48 h. A. Fluorescence image of a confluent human cFBs monolayer (F-actin in green and nuclei in blue, with 100 μm scale bar). B. Experimental design with varying plating cell density (normal: 50,000 cells per well; ½: 25,000 cells per well and ¼: 12,500 cells per well) and oxygen diffusion path through solution volume in each well (300 μL, 200 μL, 100 μL, and 50 μL per well); four biological samples per each of the 12 conditions are included. The analyzed ratiometric images at 0 h and 48 h after cell plating are shown for the 48 samples. The half-circles reflect the shape of the optical sensors; color encodes % pericellular oxygen. C. Pericellular oxygen dynamics over 48 h for the experiment in panel B, with the samples grouped by solution volume/height and color encoding the cell density. D. Pericellular oxygen dynamics as in C, with the samples grouped by cell density and color encoding used for the four solution heights. Insets are binarized images of nuclei for the different plating densities at the start of an experiment (2 h after plating) and at the 48 h endpoint, n = 4 for each of the 12 cases. Experiments have been repeated five times, with similar trend observed. E. Summary plot from independent runs for steady-state pericellular oxygen levels for normal, ½ density, and ¼ density cFBs, n = 8 for each of the 12 cases. Two-factor ANOVA revealed significant effects of cell density ( p < 0.0001) and solution height (p < 0.0001) as well as their interaction ( p = 0.0001) on the steady-state levels of pericellular oxygen in human cFB. The green box in panel E delineates the “normoxia” region for pericellular oxygen values and indicates that, for all studied conditions, cFB operated under hyperoxia.

Article Snippet: Human adult ventricular cardiac fibroblasts, cFBs (Cell Applications, Inc. 307V-75a) were thawed into a T75 flask per the manufacturer's instructions.

Techniques: Fluorescence, Diffusion-based Assay