Immunostaining of DNA in electron microscopy: an amplification and staining procedure for thin sections as alternative to gold labeling.
We describe a new electron microscopic on-section staining technique with high specificity and sensitivity for DNA-containing structures. Lowicryl HM20 sections of specimens obtained by cryofixation and freeze-substitution are incubated in a first step with a primary IgM antibody specific for double-stranded DNA. The layer of bound antibodies at the section surface is amplified in a successive step by a secondary IgG antibody. Finally, electron scattering of the antibody layer produced is enhanced by staining with a mixture of uranyl acetate and potassium permanganate. The applicability of the method is exemplified by the detection of shape and distribution of various types of bacterial and eukaryotic chromatin.
Bohrmann
B
B
Department of Physics, Pharma Research New Technologies, F. Hoffmann-La Roche Ltd., Basel, Switzerland.
Kellenberger
E
E
D000818:Animals; D002843:Chromatin / Q000648:ultrastructure; D004247:DNA / Q000648:ultrastructure*; D004269:DNA, Bacterial / Q000648:ultrastructure; D016054:DNA, Protozoan / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics / Q000648:ultrastructure; D005056:Euglena gracilis / Q000235:genetics / Q000648:ultrastructure; D007150:Immunohistochemistry; D016253:Microscopy, Immunoelectron / Q000379:methods*; D012680:Sensitivity and Specificity; D013194:Staining and Labeling / Q000379:methods; D014346:Trypanosoma brucei brucei / Q000235:genetics / Q000648:ultrastructure
D002843:Chromatin; D004269:DNA, Bacterial; D016054:DNA, Protozoan; D004247:DNA