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Mass analysis of biological macromolecular complexes by STEM
Author(s) -
Thomas Daniel,
Schultz Patrick,
Steven Alasdair C,
Wall Joseph S
Publication year - 1994
Publication title -
biology of the cell
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.543
H-Index - 85
eISSN - 1768-322X
pISSN - 0248-4900
DOI - 10.1111/j.1768-322x.1994.tb00929.x
Subject(s) - biology , scanning transmission electron microscopy , dark field microscopy , macromolecule , biophysics , molecular mass , chemical physics , biological system , nanotechnology , materials science , physics , biochemistry , transmission electron microscopy , microscopy , optics , enzyme
Scanning transmission electron microscopy (STEM) provides a superbly versatile method of measuring the masses of macromolecular complexes ranging in size from single protein subunits to large virus particles. The physical basis of the method is the elastic scattering of electrons by the component atoms of the specimen. Unstained molecules yield a dark‐field signal that is proportional to their local mass density, thus allowing direct measurements of the total mass of an individual particle, as well as of the masses of its resolved domains by integrating over appropriate regions of the image. In this review, we present an introduction to the STEM method of mass analysis from a practical standpoint, stressing the essential points of specimen preparation, as well as the scope and current limitations of the method. Its potentialities are illustrated by applications to several classes of macromolecules: isolated oligomeric proteins (the envelope glycoprotein of HIV), nucleoprotein complexes (SV40 minichromosome, transcription factor TFIIIC), membranous specimens (clathrin‐coated membranes, the VDAC channel), and viruses (vesicular stomatitis virus; herpes simplex virus). In the case of multicomponent complexes, STEM mass measurements of both the intact complex and of defined biochemical derivatives (for instance, after extraction of specific components), allow one to compile complete and precise molecular inventories. Finally, we briefly anticipate future advances that should allow even more precise and detailed mass mappings, the labelling of specific sites with heavy atom clusters, and elemental mapping based on weak inelastic signals acquired in parallel with the relatively intense dark‐field signals that have been so successfully exploited to date.