Cool snapfx camera

Embryos were transferred to glass-bottom dishes (MatTek Corp., Ashland, MA) in E3 (inverted microscope) or embedded in 0.5% E3-agarose (confocal) containing 0.02% MS222. A Leica DM IRB inverted microscope (bright-field, differential interference contrast (DIC), and fluorescence imaging) coupled to a Coolsnap fx camera (Roper Scientific) was used. A Nikon AZ100 equipped for bright-field and fluorescence imaging, coupled with Coolsnap HQ2 (Roper Scientific) using MetaVue software was used to record full size embryos. Confocal microscopy was performed with an Olympus FV10i and images and movies were processed with Fluoview software and Image J. Images were processed further using Adobe Photoshop, and time-lapse videos made with image J (see specific details in movie legends). To quantify phagocytic cells, the fluorescent pixel quantification method was used as described [69 (link)]. Graphs depict macrophage and neutrophil numbers. BioImageXD [70 (link)] was used to obtain the image in S6A Fig.

Mitochondrial fragmentation analysis was carried out as previously reported.^{29 (link)} In short, mitochondria and nucleus of the MDA-MB-231 cells were stained with Mito Tracker Red CMX Ros and DAPI, respectively. After that, the cells were treated with STE and ATE respectively for 12 h, then the cells were photographed under a monochromatic Cool SNAPFX camera (Roper Scientific, USA).

Normal human keratinocyte (NHK) cultures were established from abdominal skin samples from participants who were 20 (strain 1) and 32 (strain 2) years of age according to previously published procedures (Michopoulou et al., 2020 (link)). The cultured cells were grown in a keratinocyte growth medium supplemented with 0.15 mM CaCl₂ (KBM‐2 BulletKit, Lonza Biosciences, Basel, Switzerland). To initiate experiments, 2 × 10⁵ keratinocytes were cultured in CytoSoft® 6‐well plates, elastic moduli 2 and 32 kPa (Sigma Aldrich), for 48 h in keratinocyte basal medium (KBM)‐2. Wells were rinsed with sterile PBS and processed for RNA extraction. Cells were photographed using a Zeiss Axiovert 40 microscope equipped with a circular differential interference contrast and coupled to a Coolsnap Fx camera (Roper Scientific, Evry, France).