Abstract
BackgroundElectron tomographic analysis can be combined with the simple and rapid negative staining technique used in electron microscopy based virus diagnosis.MethodsStandard negative staining of representative examples of parapoxviruses and paramyxoviruses was combined with electron tomographic analysis.ResultsDigital sectioning of reconstructions of these viruses at a selected height demonstrated the viral ultrastructure in detail, including the characteristic diagnostic features like the surface threads on C-particles of a parapoxvirus and individual glycoproteins and the internal nucleoprotein strand of Newcastle disease virus. For both viruses, deformation and flattening were observed.ConclusionThe combination of negative staining of complex viruses with electron tomographic analysis, allows visualizing and measuring artifacts typical for negative staining. This approach allows sharp visualisation of structures in a subnanometer-thick plane, avoiding blurring due to superposition which is inherent to TEM. In selected examples, such analyses can improve diagnosis of viral agents.
Highlights
Electron tomographic analysis can be combined with the simple and rapid negative staining technique used in electron microscopy based virus diagnosis
Virus diagnosis based on morphology is hindered by the limitations of both negative staining and transmission electron microscopy (TEM)
We aimed to demonstrate that electron tomographic analysis of negatively stained complex viruses, like paramyxoviruses and parapoxviruses, allows visualizing and measuring artifacts like deformation and flattening, and yields additional and relevant structural information
Summary
Electron tomographic analysis can be combined with the simple and rapid negative staining technique used in electron microscopy based virus diagnosis. Because of the simplicity and speed of this technique, one of its major applications is the detection and identification of viruses. Such electron microscopic diagnosis is uniquely suited for the identification of infectious agents in emergent situations because of its speed, and its "open view", i.e. the ability to detect multiple viruses without the need for specific reagents [3]. Virus diagnosis based on morphology is hindered by the limitations of both negative staining and transmission electron microscopy (TEM). Limitations of TEM include the destructive effects of irradiation and blurring of fine ultrastructural details due to superpo-
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