506 059 objective

Living embryos were transferred into 2.5% methylcellulose (in E3-buffer) onto an object slide and image documentation was done with the bright-field stereomicroscope LEICA MZFLIII (Wetzlar, Germany) using the digital LEICA DFC310 FX camera, with the cold-light source KL 1500 LCD (Schott, Mainz, Germany). Video documentation for the beating hearts was performed using the LEICA DM IRB microscope with a magnification of 1.6×, a LEICA 506,059 objective, and a LEICA DFC360 FX camera (MCU II, Kappa GmbH, Germany). Images were processed with ImageJ 1.50i (National Institutes of Health, MD). Heart rate was calculated as heart beats per min. The “Zebrafs” program was used to analyze the percentage of ventricular and arterial fractional shortening.

Cardiac function in zebrafish embryos was assessed at 24 or 72 hpf. The embryos were anesthetized with 0.02% tricaine (Sigma-Aldrich, #E10521) and mounted in 2.5% methylcellulose. Video documentation was performed by recording 10 sec video frames using the LEICA DM IRB microscope with a magnification of 1.6×, a LEICA 506059 objective and a LEICA DFC360 FX camera (MCU II, Kappa GmbH, Germany). Images were processed with VirtualDub Version 1.7. Fractional shortening of heart chambers was calculated with help of the zebraFS software (www.benegfx.de).