Quorum q150t sputter coater

For the qualitative analysis of the blood clot formation, uncoated and TiC coated titanium disks were put into a 24-well dish (NuncA/S, Roskilde, Denmark) sterilized by exposing to UV for 2h, then exposed to human Whole Blood (hWB) or human Platelet-Rich Plasma (hPRP).
In the first case, the wells were filled with 3 ml of freshly drawn hWB (obtained from consenting donors) and incubated 4 min at RT. The excess liquid was then discarded and the discs were rinsed three times in a saline solution.
In the second case, the sterilized substrates were incubated with 3 ml of hPRP at 37°C, 5% CO₂ for 90 min. After the incubation time, the disks were set free of liquids by suction and rinsed in a saline solution.
After the preparation, all disks were fixed in 2.5% glutaraldehyde in 0.1M cacodilate buffer pH 7.4 for 1 h at 4°C, dehydrated through a graded series of ethanol solutions and air dried. The samples were then sputtered with 10 nm of chromium with a Quorum Q150T Sputter Coater (Quorum Technologies, Laughton, UK) and examined with a FESEM (Field Emission Scanning Electron Microscope, Auriga Zeiss).

For light microscopy, kidneys fixed in 10% neutral-buffered formalin were embedded in paraffin wax and sectioned at 5 μm. Kidney sections were stained with hematoxylin and eosin, periodic acid–Schiff, and Masson trichrome stain.
For transmission electron microscopy (TEM) and scanning electron microscopy (SEM), 1-mm³ cubes of kidney cortex were fixed in 3% glutaraldehyde and 4% formaldehyde in 0.1 M PIPES (the common name for piperazine-N,N′-bis(2-ethanesulfonic acid) buffer, pH 7.2 (minimum, 1 hour).
For TEM, specimens were then rinsed in 0.1 M PIPES buffer, postfixed in 1% buffered osmium tetroxide, rinsed in buffer, block stained in 2% aqueous uranyl acetate, dehydrated in an ethanol series, and embedded in TAAB resin (TAAB Laboratories). Gold–silver sections were cut, stained with Reynolds lead stain, and viewed on a Hitachi HT7700 transmission electron microscope.
For SEM, samples were then dehydrated through increasing strength of ethanol solutions and critical point dried using an Emitech K850 (EM Technologies LTD). Three specimens per animal were then mounted on stubs using silver paint (Agar Scientific) and sputter coated with platinum using a Quorum Q150T sputter coater (Quorum Technologies). The specimens were untimely visualized with a JEOL LSM-6010 (Jeol Ltd.).

The morphology of the prepared nanofibers and ODFs were examined using scanning electron microscopy (SEM). A 0.4 cm × 0.4 cm piece of foil on which the nanofibers were collected and an equivalent amount of ODF was adhered onto an SEM stub, using double-sided carbon tabs (Agar Scientific, Stansted, UK). The prepared stub was then given a thin coating of gold (10 nM) in a Quorum Q150T Sputter Coater (Quorum Technologies Ltd. East Sussex, UK) in an argon atmosphere. The coated stub was then transferred and imaged under FEI Quanta 200F (FEI company Ltd., Eindhoven, The Netherlands), at an acceleration voltage of 5 kV. Fiber size analysis was performed by measuring the diameter of at least 100 fibers using ImageJ software (National Institutes of Health, Bethesda, MD, USA).