On the Distortion in Dimensions Produced by Shadowing

Misra, D.N.; Gupta, Nandini; Sadhukhan, P.

doi:10.1016/b978-1-4832-2795-5.50072-1

Cited by 11 publications

(13 citation statements)

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“…The resultant plates were then examined under a binocular dissecting microscope with a calibrated scale in one eyepiece. The diameters of the chylomicrons were measured in the axis at right angles to the shadow [Mistra and Das Gupta, 1965]. This measurement was checked against polystyrene spheres of known diameter of 880±80 and 5570±100 A.…”

Section: Methodsmentioning

confidence: 99%

The Effect of Dietary Fat Load on the Size and Composition of Chylomicrons in Thoracic Duct Lymph

1968

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Chylomicrons of Sf 〉 400 and lipoproteins of Sf 12‐400 were separated by ultra‐centrifugation from samples of thoracic duct lymph collected from rabbits fed for 1 to 2 weeks previously on diets containing 5 per cent and 30 per cent corn oil or butter and from rats given a single intragastric dose of 0·5 ml. corn oil. The size of these particles after corn oil was determined by electron microscopy following fixation in osmium tetroxide. In rabbits on the low fat diet the mean diameter of chylomicrons was 960 Å with half the triglyceride in particles of diameter greater than 1400 Å; on the high fat diet the mean diameter was 1435 Å with half the triglyceride in particles of diameter greater than 2700 Å. In rats the mean chylomicron diameter was 1094 Å in the early phase of fat absorption with half the triglyceride in particles of diameter greater than 1320 Å. These figures increased to 2105 Å and 3600 Å respectively at the peak of fat absorption. The relationship between the chemical composition of these complexes and their actual size suggests that phospholipid is uniformly distributed on the surface, each phospholipid molecule occupying approximately 65 Å2. Although meaningful electron micrographs of butter chylomicrons could not be obtained, determinations of the triglyceride : phospholipid ratios indicate that, as with the corn oil‐fed animals, the chylomicrons from the rabbits fed 30 per cent butter are on the average larger than those from the animals fed 5 per cent butter.

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Section: Methodsmentioning

confidence: 99%

The Effect of Dietary Fat Load on the Size and Composition of Chylomicrons in Thoracic Duct Lymph

1968

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“…Particle sizes were measured on photographic negatives (X 40,500) or on corresponding prints (total magnification X 110,000) with an optical microcomparitor. 5 The maximum diameter of each particle was measured in the axis at right angles to its shadow (23). The diameters of 600 randomly chosen particles were measured after shadow casting of particles separated from pooled biopsy homogenates taken from six different fasting volunteers.…”

Section: Methodsmentioning

confidence: 99%

Synthesis and transport of lipoprotein particles by intestinal absorptive cells in man

Tytgat¹,

Rubin²,

Saunders³

1971

J. Clin. Invest.

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“…Previous attempts to determine the distortion of Pt replicated particles perpendicular to the shadow direction has produced metalcoated particle width distributions which are similar to log normal distributions (Hall, 1960;Misra and Das Gupta, 1965). The means of these distributions were estimated to be 5.7-7.6 nm larger than the diameter of the protein.…”

Section: Modeling How the Metal Replication Film Forms Around Particlmentioning

confidence: 99%

Quantification of particle sizes with metal replication under standard freeze‐etching conditions: A gold ball standard for calibrating shadow widths was used to measure freeze‐etched globular proteins

Ruben

1995

Microscopy Res & Technique

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The real size of platinum-carbon (Pt-C) replicated particles is not directly equivalent to either its metal-coated diameter or its shadow width. This paper describes two indirect methods, shadow widths and coated particle diameters, for determining a particle's actual size beneath a Pt-C replication film. Both produce equivalent measurements using the same standardized conditions: 2.3 nm Pt-C films deposited at a 45 degree angle on an approximately -100 degrees C surface in a 10(-6) torr vacuum. For the first method, gold balls nucleated in a partial pressure of helium and deposited on flat indirect carbon films (root mean square roughness of 0.8 nm) on 400 mesh grids were used as test particles for calibrating shadow widths as a function of particle size. The gold ball test specimens were replicated, and a distribution of Pt-C shadow widths orthogonal to the Pt-C deposition direction was measured and averaged for gold balls 1.5 +/- 0.25 nm, 2.0 +/- 0.25 nm, etc. The diameter of each gold ball was measured within the Pt-C film along with its shadow width because the Pt-C did not obscure or adhere well to the gold. The shadow width distributions for each gold size do not differ significantly from log normal. Two proteins, the lactose repressor and the mitochondrial ATPase, F1, were also used as replication test objects. Negative staining of both proteins was conducted to measure their average diameters. In the second method, a distribution of Pt-C-coated lac repressor diameters perpendicular to the shadow direction was measured. The Pt-C film thickness measured on the quartz crystal monitor was subtracted from the average metal-coated protein diameter to obtain the lac repressor's diameter. The Pt-C-coated particle diameter distributions also did not differ significantly from log normal. While doing this work it was discovered that outgassing the Pt-C electron gun greatly affected Pt-C film granularity: 19 sec produced a high contrast, granular Pt-C film, whereas 120 sec yielded a low contrast, less granular Pt-C film. Both gold balls and protein particles were subjected in separate experiments to either 19 or 120 sec of outgassing of the Pt-C gun prior to Pt-C replication. Outgassing had a profound effect on the average size of the Pt-C shadow widths on both gold and protein particles. The Pt-C gun outgassing procedure also determined the smallest replicated particle that could be resolved. The frequency of some smaller gold ball sizes detected after replication was reduced disproportionately with 19 sec vs. 120 sec outgassing. However, Pt-C gun outgassing did not affect the average measured diameter of the Pt-C-coated protein particles. The "geometric assumption" that each metal-coated particle creates a shadow width the same size as the metal-coated particle diameter was tested using a globular protein. Pt-C replication of protein particles at a 45 degree and 20 degree angle could not confirm the geometric assumption because an average shadow width was always significantly larger than its average Pt-C-coated pa...

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On the Distortion in Dimensions Produced by Shadowing

Cited by 11 publications

References 0 publications

The Effect of Dietary Fat Load on the Size and Composition of Chylomicrons in Thoracic Duct Lymph

The Effect of Dietary Fat Load on the Size and Composition of Chylomicrons in Thoracic Duct Lymph

Synthesis and transport of lipoprotein particles by intestinal absorptive cells in man

Quantification of particle sizes with metal replication under standard freeze‐etching conditions: A gold ball standard for calibrating shadow widths was used to measure freeze‐etched globular proteins

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