Carbon supported platinum and palladium alloy

BSEM imaging was used to quantify osteocyte lacunae in tibia midshaft cortical bone. Measurements of lacunar density and area were performed on images acquired by a scanning electron microscope equipped with a backscattered electron detector operated with an accelerating voltage of 20 kV, a working distance of 10.0 mm, aperture size of 30.00 μm, and 500× original magnification (SEM, Supra 55 VP, Zeiss, Center for Nanoscale Systems in Harvard University). MMA-embedded tibia sample blocks were sequentially polished with silicon carbide sandpaper of increasing grit number (240, 600, 2,400, 6,000, 8,000 grit) and alumina polishing micromesh cloth of 12,000 grit (Scientific Instrument Services Inc.). Blocks were sputter-coated with 5 nm of carbon-supported platinum and palladium alloy (Leica Microsystems Inc.) and mounted and fixed with aluminum conductive tape (Ted Pella, Inc.). BSEM images were taken at a standardized tibial midshaft area located 4.5 mm distal from the proximal tibia-fibula junction, the same standardized region used for μCT and histomorphometry analyses. Images were thresholded using default method in ImageJ 1.52d (NIH) and analyzed using Fiji software, to measure mean LcA (mm²) and LcD (#/mm²), this latter reflecting the number of lacunae per area of mineralized bone analyzed.

BSEM imaging was used to quantify osteocyte lacunae in tibia midshaft cortical bone. Measurements of lacunar density and area were performed on images acquired by a scanning electron microscope equipped with a backscattered electron detector operated with an accelerating voltage of 20 kV, a working distance of 10.0 mm, aperture size of 30.00 μm, and 500× original magnification (SEM, Supra 55 VP, Zeiss, Center for Nanoscale Systems in Harvard University). MMA-embedded tibia sample blocks were sequentially polished with silicon carbide sandpaper of increasing grit number (240, 600, 2,400, 6,000, 8,000 grit) and alumina polishing micromesh cloth of 12,000 grit (Scientific Instrument Services Inc.). Blocks were sputter-coated with 5 nm of carbon-supported platinum and palladium alloy (Leica Microsystems Inc.) and mounted and fixed with aluminum conductive tape (Ted Pella, Inc.). BSEM images were taken at a standardized tibial midshaft area located 4.5 mm distal from the proximal tibia-fibula junction, the same standardized region used for μCT and histomorphometry analyses. Images were thresholded using default method in ImageJ 1.52d (NIH) and analyzed using Fiji software, to measure mean LcA (mm²) and LcD (#/mm²), this latter reflecting the number of lacunae per area of mineralized bone analyzed.