Titre
Cryo-negative staining reduces electron-beam sensitivity of vitrified biological particles
Type
article
Institution
UNIL/CHUV/Unisanté + institutions partenaires
Périodique
Auteur(s)
De Carlo, S.
Auteure/Auteur
El-Bez, C.
Auteure/Auteur
Alvarez-Rua, C.
Auteure/Auteur
Borge, J.
Auteure/Auteur
Dubochet, J.
Auteure/Auteur
Liens vers les personnes
Liens vers les unités
ISSN
1047-8477
Statut éditorial
Publié
Date de publication
2002-06
Volume
138
Numéro
3
Première page
216
Dernière page/numéro d’article
26
Notes
Journal Article --- Old month value: Jun
Résumé
Beam damage is the main resolution-limiting factor when biological particles are observed by cryoelectron microscopy in a thin vitrified solution film. Furthermore, the low contrast of the specimen frequently makes observation difficult and limits the possibility of image processing. Cryo-negative staining, in which the particles are vitrified in a thin layer of concentrated ammonium molybdate solution, makes it possible to visualize the particles with a much better signal-to-noise ratio (SNR) while keeping the specimen in a good state of preservation. We have observed the Escherichia coli GroEL chaperonin, prepared in a native vitrified solution and by cryo-negative staining after electron exposure from 1000 to 3000e(-)/nm(2). We have compared the resulting three-dimensional models obtained from these different conditions and have tested their fit with the atomic model of the protein subunit obtained from X-ray crystallography. It is found that, down to 1.5-nm resolution, the particles appear to be faithfully represented in the cryo-negatively stained preparation, but there is an approximately 10-fold increase of SNR compared with the native vitrified preparation. Furthermore, for the same range of irradiation and down to the same resolution, the particles seem unaffected by beam damage, whereas the damage is severe in the native vitrified particles.
Sujets
PID Serval
serval:BIB_625F3955BFFC
PMID
Date de création
2008-01-24T09:25:37.891Z
Date de création dans IRIS
2025-05-20T23:22:42Z