Determination of protein and nucleic acid concentration and localization of specific elements by scanning transmission electron microscopy.
Elastic and inelastic dark-field micrographs simultaneously recorded by the scanning transmission electron microscope (STEM) provide structural and quantitative chemical information at a spatial resolution of some nanometers. Simultaneous acquisition is essential as it warrants: (i) the geometrical identity of picture elements (pixels) in these micrographs, and (ii) the identity of recording conditions (focus, electron dose) for the elastic and inelastic images. Suitable off-line processing of such multichannel images allows the concentration of protein or nucleic acid within embedding material to be evaluated. In addition, location and number of chemical elements such as phosphorus can be determined. Theoretical sensitivity is a few phosphorus atoms in a volume of approximately 1000 nm3 (not demonstrated in this paper). Quantitative electron microscopy of this kind, however, requires the influence of mass loss as well as plural scattering to be correctly accounted for. Experimental conditions that provide a sound basis for concentration determination as well as element mapping are discussed.
Reichelt
R
R
M.E. Müller-Institute for High Resolution Electron Microscopy, University of Basel, Switzerland.
Engel
A
A
D000818:Animals; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D009696:Nucleic Acids / Q000032:analysis*; D011506:Proteins / Q000032:analysis*
D009696:Nucleic Acids; D011506:Proteins