Cellobserver z 1

Pupae were imaged whole with a Leica Fluorescence Stereo Microscope and Leica Application Suite X (LasX, Version 3.5.2.18963).
Confocal fluorescent images were taken with a Leica TCS SP8 with an HC PL APO CS2 63×/1.4 oil objective and Leica Application Suite X (LasX, Version 3.5.2.18963). Live imaging of macrophage cultures was performed using a Zeiss CellObserver Z.1 with a Yokogawa CSU-X1 spinning disk scanning unit and an Axiocam MRm CCD camera (6.45 µm×6.45 µm) and ZenBlue 2.5 software. Ablation experiments were performed using the UV ablation system DL-355/14 from Rapp OptoElectronics, as reported previously (Lehne et al., 2022 (link)).

Structure illumination microscopy images were taken with an ELYRA S.1 microscope (Cell Observer SD, 63×/1.4 oil-immersion objective). Confocal fluorescence images were taken using a Leica TCS SP8 with an HC PL APO CS2 63×/1.4 oil objective. Live imaging of macrophage cultures was performed using a Zeiss CellObserver Z.1 with a Yokogawa CSU-X1 spinning disc scanning unit and an Axiocam MRm CCD camera (6.45 µm×6.45 µm). Ablation experiments were done using a 355 nm pulsed UV laser (Rapp, Optoelectronics), as reported previously (Sander et al., 2013 (link); Rüder et al., 2018 (link)).

Confocal images were taken with a Leica TCS SP8 with an HC PL APO CS2 ×63/1.4 oil objective and Leica Application Suite X (LasX) software. Structure illumination microscopic images were taken with a Zeiss ELYRA PS1 Microscope with a ×63/1.4 oil objective. Live imaging of macrophages was performed using a Zeiss CellObserver Z.1 with a Yokogawa CSU-X1 spinning disk scanning unit and an Axiocam MRm CCD camera (6.45 µm × 6.45 µm) and ZenBlue 2.5 software. Laser ablation of single cells was done using the UV laser ablation system DL-355/14 from Rapp OptoElectronics. Imaging of fixed B16-F1 cells was performed with an Olympus XI-81 inverted microscope equipped with an UPlan FI ×100/1.30NA oil immersion objective or a Zeiss LSM980 confocal microscope equipped with a Plan-Neofluar ×63/1.45NA oil immersion objective using 488 nm and 561 nm laser lines. Fluorescence intensities of phalloidin-stained lamellipodia were quantified from 8-bit images captured at identical settings using ImageJ software after background subtraction. Relative mean pixel intensities in lamellipodial regions of interest are shown as whiskers-box plots including all data points. The numbers of microspikes were manually counted.