title: Examination of electron stains as a substitute for uranyl acetate for the ultrathin sections of bacterial cells.
abstract: Electron staining reagents were examined to find a possible substitute for uranyl acetate (UA) in electron microscopy of bacterial ultrathin sections. Four kinds of stains, platinum blue (Pt-blue), oolong tea extract (OTE), potassium permanganate (KMnO(4)) and phosphotungstic acid (PTA), were examined in comparison with UA either with or without post-staining with lead citrate (Pb). Electron microscopy was performed on sections from Spurr-embedded cells of a Gram-positive bacterium, Bacillus cereus NBRC 13597, and a Gram-negative bacterium, Escherichia coli NBRC 3301. Both Pt-blue and OTE showed staining similar to each other and to that of double staining with UA and Pb in B. cereus, while in E. coli the cytoplasmic membrane appeared less dense when compared with UA and Pb. KMnO(4) stained excessively to some extent, but showed images of the best contrast in the cytoplasmic membrane comparable with UA and Pb among the four reagents. PTA could stain the peptidoglycan layer but gave images of low quality for both bacteria. This study demonstrated that none of the reagents examined showed staining results of the same quality or better than the conventional method with UA and Pb. However, stains of Pt-blue, OTE and KMnO(4) could possibly be an alternative candidate for the UA according to the structure in question.
authors.affiliation: Biological Resource Center (NBRC), Department of Biotechnology, National Institute of Technology and Evaluation, 2-5-8 Kazusakamatari, Kisarazu, Chiba 292-0818, Japan. yamaguchi-kaoru@nite.go.jp
authors.forename: Kaoru
authors.initials: K
authors.lastname: Yamaguchi
authors.forename: Ken-ichiro
authors.initials: K
authors.lastname: Suzuki
authors.forename: Kenji
authors.initials: K
authors.lastname: Tanaka
mesh_terms: D001409:Bacillus cereus / Q000648:ultrastructure*; D004583:Electrons; D004926:Escherichia coli / Q000648:ultrastructure*; D017110:Freeze Substitution; D007854:Lead / Q000378:metabolism; D008854:Microscopy, Electron / Q000379:methods*; D008867:Microtomy; D009942:Organometallic Compounds; D010772:Phosphotungstic Acid; D010984:Platinum; D011196:Potassium Permanganate; D013194:Staining and Labeling / Q000379:methods*; D016610:Tissue Embedding
chemical_list: D009942:Organometallic Compounds; D011196:Potassium Permanganate; D010772:Phosphotungstic Acid; C005460:uranyl acetate; D007854:Lead; D010984:Platinum