Fv1200mpe laser confocal microscope

Triple TOF™ 5600 + high resolution tandem mass spectrometry (AB SCIEX, Redwood City, CA);
Ultra-high performance liquid chromatography system (Shimazhu 20A, Shimadzu Corporation,
Tokyo, Japan); D3024R bench refrigerated centrifuge (Beijing Dalong Co., Ltd., Beijing,
China); EYELLA N1100 rotary evaporator (Tokyo Rikakikai Co., Ltd., Tokyo, Japan);
Ultrasonic crushing instrument (Wuxi Voshin instruments Manufacturing Co., Ltd., Huishan,
China); Nano-ZS particle size tester (Malvern Instruments, Worcestershire, UK);
Ultimate3000 high performance liquid chromatography (Thermo Fisher Scientific, Waltham,
MA); pectraMax Plus384 Molecular Devices (Molecular Devices, Silicon Valley, CA); HF240
cell culture box (Shanghai Lishen Scientific Instrument Co., Ltd., Shanghai, China);
SW-CJ-2FD clean bench (Suzhou Antai Airtech Co., Ltd, Beijing, China); T9S dual-beam
ultraviolet-visible spectrophotometer (Beijing Persee General Instrument Co., Ltd,
Beijing, China); FV1200MPE laser confocal microscope (Olympus, Tokyo, Japan).

NIH/3T3 fibroblasts were seeded on glass-bottom MatTek dishes (MatTek Corp., USA) coated with rat tail type I collagen, as described [16 (link),17 (link)]. Adhered cells were treated with vehicle or TGF-β1 (5 ng /mL) for 48 hours, as described above. For life cell imaging of cell nuclei and intracellular mitochondria, cells were loaded with cell-permeable nuclear-selective fluorescent dye Hoechst 33342 (Molecular Probes, USA), and red fluorescing mitochondrial membrane potential sensitive dye tetramethylrhodamine (TMRM) (Molecular Probes, USA). The fluorescent images of stained cells were obtained using an Olympus FV1200 MPE laser confocal microscope (Olympus, USA) equipped with a long working distance dry UPLSAPO 40X/0.95 lens with corrective collar and appropriate excitation laser lines with corresponding dichroic cubes (Hoechst 33342; excitation 405 nm/emission 470 and TMRM; excitation 569 nm/emission 590 nm).