Agar sputter coater b7340

Markings were created at 2, 4, and 6 mm from the tip of each root horizontally cut with a diamond disc using water as a coolant. A cone of Gutta-percha was put into the root canal to cover the canal walls and reduce debris from the sectioning. The teeth were coated with gold-palladium sputter (Agar Sputter Coater B7340; Agar Scientific Ltd., Stanton, UK) to prepare a scanning electron microscope (JEOL Ltd., Tokyo, Japan) at 1000× magnification, and photomicrographs were taken and evaluated. Two independent examiners were trained to determine the presence of a smear layer according to Torabinejad’s et al. [17 (link)] description, where a score of zero has no smear layer on the surface of the root canal, and all dentinal tubules are clean and open, and score one has a moderate smear layer on the surface of the root canal. However, there is debris in the dentinal tubules, and a thick smearing layer covers the root canal surface and dentinal tubules in score two. The Kappa (k) statistic was used to assess inter- and intra-observer agreement using the following interpretation paradigm: poor (0.20), fair (0.21–0.40), moderate (0.41–0.60), good (0.61–0.80), and excellent (0.81–1.00) [18 (link)].

Native, pre-digested and fermented wheat bran samples were visualized using cryo-SEM and desktop SEM. For the purpose of SEM microscopy, the bran samples were chemically dried with hexamethyldisilazane (HMDS) as described by Araujo et al. (2003) (link). After complete evaporation of the HMDS, samples were mounted on an aluminum pin (diameter: 12 mm) using double sided carbon tape and subsequently gold sputtered for 45 s at 30 mA (Agar Sputter Coater B7340, Agar Scientific, UK). Images were collected using a Phenom Pro X SEM microscope (Phenom-World B.V., the Netherlands) with a beam intensity of 10 keV.
As an alternative to SEM microscopy, samples were also visualized via cryo-SEM using a Jeol JSM 7100F scanning electron microscope (JEOL Ltd, Tokyo, Japan). A small amount of wheat bran was placed on a sticky carbon surface mounted on an aluminium stub, vitrified in a nitrogen slush and transferred under vacuum conditions into the cryo-preparation chamber (PP3010T Cryo-SEM Preparation System; Quorum Technologies, Lewes, UK) conditioned at −140 °C. Subsequently, the sample was sublimated for 20 min at −70 °C to remove frost artefacts, sputter-coated with platinum using argon gas, transferred to the SEM stage at −140 °C and electron beam targeted at 3 keV.