Dark-field electron microscopy of platelet adhesive macromolecules.
Application of these methods to a number of macromolecular species shows, in general, that features which are only occasionally or marginally detected in bright field may be visualized clearly and consistently through the use of dark-field illumination. Given the minimal average thicknesses of contrasting metal required for dark-field examination (ca. less than 10(-7) g/cm2, or less than 0.4 nm thickness), distortions of molecular features by accumulation of coating metal, although not avoided altogether, are certainly minimized. Thus, application of minimal metal coating in conjunction with high-resolution dark-field electron microscopy should facilitate solution of structural problems in molecular biochemistry generally. The method is of particular current interest in the specific area of coagulation and adhesive protein conformation. The advantages of metal coating macromolecules for dark-field observation are summarized in Table I. In the case of macromolecules or macromolecular complexes which are of limited width (5 nm or less) even though extended lengthwise, contrast available for visibility tends to be limiting. It is for such structures that dark field offers a productive alternative. The dark-field method seems promising for study of certain macromolecular preparations of adhesive proteins vis-à-vis isolation of receptor loci. These include the complex between fibrinogen and GPIIb/IIIa; the complex between GPIb and von Willebrand protein, and the proposed complex between thrombospondin and fibrinogen. The use, for relating epitopic sites to topographical position, of Fab fragments directed against specific epitopes on coagulation macromolecules (and/or their complexes) of otherwise well-defined structure may also be extended by means of the dark-field approach.
Slayter
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D005340:Fibrinogen / Q000648:ultrastructure*; D006023:Glycoproteins / Q000648:ultrastructure*; D006801:Humans; D007202:Indicators and Reagents; D046911:Macromolecular Substances; D008854:Microscopy, Electron / Q000379:methods; D010973:Platelet Adhesiveness; D019699:Thrombospondins; D014414:Tungsten; D014841:von Willebrand Factor / Q000648:ultrastructure*
D006023:Glycoproteins; D007202:Indicators and Reagents; D046911:Macromolecular Substances; D019699:Thrombospondins; D014841:von Willebrand Factor; D005340:Fibrinogen; D014414:Tungsten