For almost four decades, the scanning transmission electron microscope (STEM) has made significant contributions to structural biology by providing accurate determinations of the molecular masses of large protein assemblies that have arbitrary shapes and sizes. Nevertheless, STEM mass mapping has been implemented in very few laboratories, most of which have employed cold-field emission gun (FEG) electron sources operating at acceleration voltages of 100 kV and lower. Here we show that a 300 kV commercial transmission electron microscope (TEM) equipped with a thermally assisted Shottky FEG can also provide accurate STEM mass measurements. Using the elastic-scattering cross sections from the NIST database, we show that the measured absolute mass values for tobacco mosaic virus and limpet hemocyanin didecamers agree with the known values to within better than 10%. We find that the measured molecular weight of the hemocyanin assemblies agrees with the known value to within 3%. FEG TEMs operating at intermediate voltages (200-400 kV) are becoming common tools for determining the structure of frozen hydrated protein assemblies. The ability to perform mass determination with the same instrument can provide important complementary information about the numbers of subunits comprising the protein assemblies whose structures are being studied.
| Sarin, Hemant; Kanevsky, Ariel S; Wu, Haitao et al. (2009) Physiologic upper limit of pore size in the blood-tumor barrier of malignant solid tumors. J Transl Med 7:51 |
