Leica mz16f microscope

All zebrafish work was done in the Tg(fli1:GFP) line [42 (link)] using an Animal Study Protocol (G-05–5 to RS) approved by the National Human Genome Research Institute’s Animal Care and Use Committee. Embryos were generated from a breeding stock, maintained at 28 °C in E3 water, and treated with 0.3 mg/mL PTU (N-Phenylthiourea) from 24 h post-fertilization (hpf) to suppress pigmentation. At 48 hpf, embryos were manually dechorionated, and anesthetized with 0.16 mg/ml tricaine (Syndel, Ferndale, WA). Seventy-five freshly thawed passage 4 HPSLFs or NLFs were suspended in 4 nL of 3 mg/mL Matrigel (Corning, Corning, NY) and were injected into the yolk sac of 48 h post-fertilization zebrafish embryos using a microinjector (World Precision Instruments, Sarasota, FL). Zebrafish embryos were screened for CM-Dil signal using a Leica MZ16F microscope (Leica, Wetzlar, Germany) 2 h after injection. Tile images from zebrafish mounted on glass-bottom microwell dishes (MatTek, Ashland, MA) in 0.8% low-melt agarose (Sigma-Aldrich, St. Louis, MO) containing 0.16 mg/mL tricaine were obtained at 24 and 48 h after injection using a Zeiss LSM 810 confocal microscope (Zeiss, Oberkochen, Germany). At least 20 zebrafish embryos were analyzed for experiments performed in triplicate using cells from 2 different patients with HPS-1 pulmonary fibrosis.