By M. A. Hayat
Uncomplicated strategies For Transmission Electron Microscopy
summary: uncomplicated recommendations For Transmission Electron Microscopy
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9% saline to flow through. Any leaks or opened side vessels are clamped. 4) is allowed to flow through the vessel. The fixative can be dyed with methylene blue to monitor the proper flow of the fixative. The saline bath in which the section of the aorta has been immersed is replaced by 3 % glutaraldehyde. The fixative can be recovered as it flows out of the system and poured back into the container for recirculation so as to maintain the desired height of the column (h in Fig. 1) of the fixative.
If possible, rinsing should be accomplished in the same buffer as that used with the fixative, to minimize differences in effective osmolality. Two or three rinses in buffer for a total period of 20 min are adequate; longer durations are required for removing the fixative before incubation for enzyme cytochemistry. Dehydration is necessary because commonly used embedding resins are im miscible with water. Therefore all the free water from the fixed and rinsed specimens must be replaced with a suitable organic solvent prior to embedding.
Several equally effective anesthetics are available for animals of different body weights. , rat and guinea pig) can be anesthetized with a volatile anesthetic such as ether or halothane. However, barbital anesthesia is preferred over ether, because incomplete perfusion of the medulla has been shown to occur in animals anesthetized with ether (Bohman, 1974). For large animals an intravenous or intraperitoneal injection with pentobarbital (Nembutal), Inactin, urethane, or chloral hydrate is used.