Semithin sections and transmission electron microscopy examination
For transmission electron microscopic investigation (TEM), parts from the eyelids were cut into small pieces (1 mm each) and were fixed in a cacodylate-buffered solution of 5% glutaraldehyde for 2 h, then washed several times in the same buffer for 1hr at pH = 7.2, and then, the specimens were post-fixed in a cacodylate buffer 1% osmium tetraoxide for 2 h at 4°C. Specimens were washed several times in the second step, followed by dehydration in a graded series of alcohol. The specimens were embedded in epoxy resin; they were treated for semi-thin sectioning at 1 mm thickness and stained with toluidine blue for light microscopic examination. Photographs were taken with an Olympus camera model DP74 connected with an Olympus microscope model BX43.
Ultrathin sections of 70 nm thickness were cut by Ultratome VRV (LKB, Bromma, Sweden), and stained with uranyl acetate and lead citrate (Reynolds, 1963), then were examined by JEOL 100CX II transmission electron microscope at the Electron Microscopy Unit of Assiut University, Egypt.