TY - JOUR
T1 - Radial mass density functions of vitrified helical specimens determined by scanning transmission electron microscopy
T2 - their potential use as substitutes for equatorial data
AU - Trachtenberg, S.
AU - Leonard, K. R.
AU - Tichelaar, W.
PY - 1992/11/1
Y1 - 1992/11/1
N2 - Using STEM dark field images, we have determined linear mass densities and radial density profiles of vitrified helical particles. The samples studied are: TMV, RNA-free helical polymers of TMV coat protein (TMV-P), Salmonella typhimurium bacterial flagellar filaments and Escherichia coli pili. The difference between the profiles obtained for TMV and TMV-P shows a maximum at a radius of about 4 nm, corresponding to the RNA in TMV. Of the peaks that are resolved in X-ray diffraction analysis we can resolve the ones for TMV at radii of ~4.2 and ~6.7 nm and a shoulder at ~7.8 nm. Density peaks in bacterial flagellar filaments appear at radii of ~4.2, ~6.5, ~8.5, and ~10.5 nm. Accurate mass data can be obtained if the filaments are embedded in ice layers of uniform thickness; their diameters need to be similar to that of the mass standard (TMV) when these data are measured in a comparative manner. Ice layers are often not uniform, and thickness variations are well revealed in STEM dark field. The signal-to-noise ratio and contrast for the transverse projections are lower than those measured for freeze-dried specimens: half an order and one order of magnitude, respectively. The thinnest uniformly thick ice layer still containing a single layer of particles is ~10-15 nm thicker than the particles. Radial mass density functions that are directly determined in STEM may have a potential use as substitutes for the unreliable equatorial data in helical reconstructions of TEM bright field images of vitrified specimens.
AB - Using STEM dark field images, we have determined linear mass densities and radial density profiles of vitrified helical particles. The samples studied are: TMV, RNA-free helical polymers of TMV coat protein (TMV-P), Salmonella typhimurium bacterial flagellar filaments and Escherichia coli pili. The difference between the profiles obtained for TMV and TMV-P shows a maximum at a radius of about 4 nm, corresponding to the RNA in TMV. Of the peaks that are resolved in X-ray diffraction analysis we can resolve the ones for TMV at radii of ~4.2 and ~6.7 nm and a shoulder at ~7.8 nm. Density peaks in bacterial flagellar filaments appear at radii of ~4.2, ~6.5, ~8.5, and ~10.5 nm. Accurate mass data can be obtained if the filaments are embedded in ice layers of uniform thickness; their diameters need to be similar to that of the mass standard (TMV) when these data are measured in a comparative manner. Ice layers are often not uniform, and thickness variations are well revealed in STEM dark field. The signal-to-noise ratio and contrast for the transverse projections are lower than those measured for freeze-dried specimens: half an order and one order of magnitude, respectively. The thinnest uniformly thick ice layer still containing a single layer of particles is ~10-15 nm thicker than the particles. Radial mass density functions that are directly determined in STEM may have a potential use as substitutes for the unreliable equatorial data in helical reconstructions of TEM bright field images of vitrified specimens.
UR - http://www.scopus.com/inward/record.url?scp=0026953527&partnerID=8YFLogxK
U2 - 10.1016/0304-3991(92)90142-7
DO - 10.1016/0304-3991(92)90142-7
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C2 - 1362014
AN - SCOPUS:0026953527
SN - 0304-3991
VL - 45
SP - 307
EP - 321
JO - Ultramicroscopy
JF - Ultramicroscopy
IS - 3-4
ER -