Diagnostic virology in veterinary pathology: techniques for negative staining

Abstract

Negative staining of virus suspensions is the quickest, easiest diagnostic technique currently available to the electron microscopist. Virus sampling and staining from directly collected fluids is inherently rapid, simple, and straightforward. Virions in a sample can be stained directly on a formvar coated grid then examined in a transmission electron microscope (TEM).The collection, detection, and identification of enteric viruses is more involved. Intestinal contents are often presented in a variety of states, from clear fluid to solid. In a diagnostic setting, a routinely reliable protocol is a necessity for fast, accurate results. Several authors have recommended various techniques for clarifying virus preparations from fecal samples. However, these methods do not inactivate pathogens in the samples. Moreover, a recent report indicates that technical staining parameters of virus staining could influence virus particle integrity. The routine use of 10% neutral buffered formalin (NBF) fixation prior to concentrating and staining virus samples both stabilizes and inactivates the majority of viruses encountered in a diagnostic setting.