Ultrathin (1 nm) vertically shadowed platinum-carbon replicas for imaging individual molecules in freeze-etched biological DNA and material science metal and plastic specimens.
Single molecule resolution in beam-sensitive, uncoated, noncrystalline materials has heretofore not been possible except in thin (less than or equal to 150 A) platinum-carbon (Pt-C) replicas, which are resistant to electron beam destruction. Previously, the granularity of metal film replicas limited their resolution to greater than or equal to 20 A. This paper demonstrates that Pt-C film granularity and resolution are a function of the method of replication and other controllable factors. Low-angle 20 degrees rotary, 45 degrees unidirectional, and vertical 9.7 +/- 1 A Pt-C films deposited on mica under the same conditions were compared. Vertical replication had a 5 A granularity, the highest resolution, and evenly coated the whole surface. A 45 degrees replication had a 9.5 A granularity, a slightly poorer resolution, and a discontinuous surface coating. The use of 20 degrees rotary replication proved to be unsuitable for high-resolution imaging, with 20-25 A granularity and resolution two to three times poorer. Vertical and 45 degrees Pt-C replicas can visualize the deep-etched DNA helix and the 13.3 A 3(2) helix of pectin in a gel. The DNA double helix, the complex structures of sol-gel glasses, Immobilon filters (polyvinylidene fluoride), a polymethacrylate plastic, the metal oxide surfaces of 440c stainless steel, and aluminum are illustrated. This high-resolution vertical Pt-C replica technique can image in the context of solutions, gels, or solids, single molecular chains 3-7 A wide, their associations, and their conformation. Included in the present article are first time descriptions for removing replicas from metals and plastics and for making high-magnification photographic prints of normal contrast using a reversal rephotographic process.
Ruben
G C
GC
Department of Biological Sciences, Dartmouth College, Hanover, New Hampshire 03755.
D002244:Carbon; D004247:DNA / Q000648:ultrastructure*; D005614:Freeze Fracturing / Q000379:methods*; D008670:Metals; D008854:Microscopy, Electron; D010969:Plastics; D010984:Platinum; D013048:Specimen Handling
D008670:Metals; D010969:Plastics; D010984:Platinum; D002244:Carbon; D004247:DNA