Specimens were imaged using a Hitachi TM3030 SEM (Hitachi Ltd., Tokyo, Japan) at 15 kV. Surfaces were scanned at 100× magnification, at 1000× magnification and 2000× magnification (n = 5). Under vacuum conditions, an electron beam was focused on the surface of the sample. In addition to preparing specimens through fixation and dehydration described previously [4 (link)], for SEM, specimens underwent sputter coating. Prior to imaging samples using SEM, specimens were mounted on an SEM stub with conductive double-sided tape and were sputter coated with gold at 2.5 kV using an Agar automatic sputter coater (Agar Scientific, Elektron, Technology UK Ltd, Essex, UK) for 30 s at 20 mA, followed by a further 30 s at 30 mA, to ensure an even covering of 150–200 Å [15 ].
For 3D surface reconstruction, four two-dimensional (2D) surface scans are stitched together using 3D-VIEW software (Hitachi Ltd., Tokyo, Japan) [18 ]. Ra was measured from the 3D reconstructed surfaces.
For 3D surface reconstruction, four two-dimensional (2D) surface scans are stitched together using 3D-VIEW software (Hitachi Ltd., Tokyo, Japan) [18 ]. Ra was measured from the 3D reconstructed surfaces.