Jsm 6340f scanning electron microscope

Microbial cells for scanning electron microscopy were collected from the bioreactors at the same time as the samples for DNA extraction described above. The cells were fixed with 2% paraformaldehyde and 2% glutaraldehyde at 4°C overnight, and then with 1% tannic acid at 4°C for 2 h. The cells were then rinsed four times with 0.1 mM cacodylate buffer and fixed with 2% OsO₄ at 4°C for 3 h. After the cells were dehydrated using a graded series of ethanol solutions, they were substituted into 100% tert-butyl alcohol and vacuumed dried. The resulting specimen was coated with a thin layer (50 nm) of osmium. The specimen was observed with a JSM-6340F scanning electron microscope (Jeol, Tokyo, Japan).

Sperms prepared from the cauda epididymis and suspended as above were fixed with an equal volume of 2% PFA and 2% GA in 0.1 M cacodylate buffer. Thereafter, they were fixed with 1% GA in 0.1 M cacodylate buffer, pH 7.4, at 4°C overnight. The samples were further fixed with 1% tannic acid in 0.1 M cacodylate buffer, pH 7.4, at 4°C for 1 h. After the fixation, the samples were washed four times with 0.1 M cacodylate buffer for 30 min each, followed by postfixation with 2% OsO₄ in 0.1 M cacodylate buffer at 4°C for 2 h. The samples were dehydrated through a series of graded ethanol (50%, 70%, 90%, 100%). The samples were substituted into tert-butyl alcohol at room temperature and then frozen. The frozen samples were vacuum dried. After drying, the samples were coated with a thin layer (30 nm) of osmium by using an osmium plasma coater (NL-OPC80NS, Nippon Laser & Electronics Laboratory, Nagoya, Japan). The samples were observed by a JSM-6340F scanning electron microscope (JEOL, Akishima, Japan) at an acceleration voltage of 5.0 kV.