Saliva Specimen Preparation for Electron Microscopy

Saliva samples were fixed with glutaraldehyde (2.5%) in a 0.1 M sodium cacodylate buffer for 12 h at 4°C. Saliva samples were deposited in Greiner Bio‐One 96‐well single‐break strip microplates (Greiner Bio‐One) coated with poly‐L lysine to retain the material. Resin embedding of the microplates was then performed as previously described [14 (link)]. Resin embedding was microwave‐assisted with PELCO BiowavePro+ (Ted Pella). Samples were washed with a mixture of 0.2 M saccharose/0.1 M sodium cacodylate and post‐fixed with 1% OsO₄ diluted in 0.2 M potassium hexa‐cyanoferrate (III)/0.1 251 M sodium cacodylate buffer. After being washed with distilled water, samples were gradually dehydrated by successive baths containing 30% to 100% ethanol. Substitution with Epon resin was achieved by incubations with 25%–100% Epon resin, and samples were placed in a polymerization chamber. Resin microwave‐curing was performed for a total of 2 h. After curing, the resin blocks were manually trimmed with a razor blade and the bases of the dishes were detached by cold shock via immersion in liquid nitrogen for 20 s. Resin blocks were placed in a UC7 ultramicrotome (Leica), trimmed to pyramids, and ultrathin 100 nm sections were cut and placed on HR25 300 Mesh Copper/Rhodium grids (TAAB).