Particle Counting and Detection

Author(s):  
Frances L. Ball ◽  
W. W. Harris

The limit of detection of viral particles with UV absorption is 1010 particles per milliliter and that of the routinely employed electron microscopy technique is 107 to 108 particles per milliliter so that low titre virus preparations such as hepatitis serum must be concentrated severalfold if these physical methods are to be used.The sensitivity of the electron microscope can be increased if all the particles in a given volume of sample can be sedimented onto one microscope grid and a study of this has been made using polio virus which titred 4 X 1012 by light scattering and 6 X 1011 by sedimentation. The number of micrographs and the necessary magnification to detect 105 particles per milliliter were calculated.

2019 ◽  
Vol 23 (3) ◽  
pp. 337-342 ◽  
Author(s):  
B. N. Zaitsev ◽  
O. S. Taranov ◽  
N. B. Rudometova ◽  
N. S. Shcherbakova ◽  
A. A. Ilyichev ◽  
...  

Viruses can infect all types of life forms, from animals and plants to microorganisms, including bacteria and archaea. When studying samples containing viruses, one confronts an unavoidable question of the quantitative determination of viral particles in the sample. One of the simplest and efficient approaches to quantitative determination of viral particles in preparation includes the use of electron microscopy; however, a high detection threshold is a significant limitation of this method (107 particles per ml). Usually, such sensitivity is insufficient and can result in error diagnosis. This study aims to develop a method making it possible to detect the number of viral particles more precisely and work with samples in which the concentration of particles is lower than 107/ml. The method includes a concentration of viral particles on the polyethersulfone membrane applied in centrifugal concentrators and subsequent calculation using an electron microscope. We selected env-pseudoviruses using a lentiviral system making it possible to obtain standardized samples of virus-like particles that are safer than a live virus. Suspension of viral particles (a volume of 20 ml) was placed into the centrifugal concentrator and centrifuged. After that, we took a membrane out of the centrifugal concentrator and evaluated the number of particles on the ultrathin section using an electron microscope. The number of viral particles on the whole surface of the filter (a square of 4 сm2) was 4×107 virions, the initial concentration of pseudoviruses in the sample was 2×106 per 1 ml (4×107 particles per 20 ml). As a result, the developed method enables one to evade the major disadvantage of quantitative determination of viruses using electron microscopy regarding a high detection threshold (concentration of particles 107/ml). Furthermore, the centrifugal concentrator makes it possible to sequentially drift a considerable volume of the suspension through the filter resulting in enhancement of test sensitivity. The developed approach results in increased sensitivity, accuracy, and reproducibility of quantitative analysis of various samples containing animal, plant or human viruses using electron microscopy.


Author(s):  
Linda M. Stannard ◽  
Margaret Lennon

Burnupena cincta and Fusus verruculatus are two whelks which inhabit the intertidal zones of the Cape Peninsula shore. Their respiratory pigments, or haemocyanins, are morphologically similar in structure (Figs. 1 and 2) and appear in the electron microscope as short cylindrical rods about 34 nm in diameter and 36 nm high. Viewed side-on the molecules show regular banding suggesting a structure composed of six equidistant rings of sub-units. Occasionally the particles have the appearance of possessing a central “belt” in the position of the 3rd and 4th rows of sub-units. End-on views of the haemocyanin molecules show a circular contour with a dense outer ring and a less dense inner ring in which 10 definite sub-units may frequently be distinguished. A number of molecules display an extra central inner component which appears either as a diffuse plug or as a discrete ring-shaped core ± 8 nm in diameter.


Author(s):  
O.C. de Hodgins ◽  
K. R. Lawless ◽  
R. Anderson

Commercial polyimide films have shown to be homogeneous on a scale of 5 to 200 nm. The observation of Skybond (SKB) 705 and PI5878 was carried out by using a Philips 400, 120 KeV STEM. The objective was to elucidate the structural features of the polymeric samples. The specimens were spun and cured at stepped temperatures in an inert atmosphere and cooled slowly for eight hours. TEM micrographs showed heterogeneities (or nodular structures) generally on a scale of 100 nm for PI5878 and approximately 40 nm for SKB 705, present in large volume fractions of both specimens. See Figures 1 and 2. It is possible that the nodulus observed may be associated with surface effects and the structure of the polymers be regarded as random amorphous arrays. Diffraction patterns of the matrix and the nodular areas showed different amorphous ring patterns in both materials. The specimens were viewed in both bright and dark fields using a high resolution electron microscope which provided magnifications of 100,000X or more on the photographic plates if desired.


Author(s):  
J. D. Hutchison

When the transmission electron microscope was commercially introduced a few years ago, it was heralded as one of the most significant aids to medical research of the century. It continues to occupy that niche; however, the scanning electron microscope is gaining rapidly in relative importance as it fills the gap between conventional optical microscopy and transmission electron microscopy.IBM Boulder is conducting three major programs in cooperation with the Colorado School of Medicine. These are the study of the mechanism of failure of the prosthetic heart valve, the study of the ultrastructure of lung tissue, and the definition of the function of the cilia of the ventricular ependyma of the brain.


Author(s):  
Lawrence W. Ortiz ◽  
Bonnie L. Isom

A procedure is described for the quantitative transfer of fibers and particulates collected on membrane filters to electron microscope (EM) grids. Various Millipore MF filters (Millipore AA, HA, GS, and VM; 0.8, 0.45, 0.22 and 0.05 μm mean pore size) have been used with success. Observed particle losses have not been size dependent and have not exceeded 10%. With fibers (glass or asbestos) as the collected media this observed loss is approximately 3%.


Author(s):  
E. U. Lee ◽  
P. A. Garner ◽  
J. S. Owens

Evidence for ordering (1-6) of interstitial impurities (O and C) has been obtained in b.c.c. metals, such as niobium and tantalum. In this paper we report the atomic and microstructural changes in an oxygenated c.p.h. metal (alpha titanium) as observed by transmission electron microscopy and diffraction.Oxygen was introduced into zone-refined iodide titanium sheets of 0.005 in. thickness in an atmosphere of oxygen and argon at 650°C, homogenized at 800°C and furnace-cooled in argon. Subsequently, thin foils were prepared by electrolytic polishing and examined in a JEM-7 electron microscope, operated at 100 KV.


Author(s):  
J. L. Farrant ◽  
J. D. McLean

For electron microscope techniques such as ferritin-labeled antibody staining it would be advantageous to have available a simple means of thin sectioning biological material without subjecting it to lipid solvents, impregnation with plastic monomers and their subsequent polymerization. With this aim in view we have re-examined the use of protein as an embedding medium. Gelatin which has been used in the past is not very satisfactory both because of its fibrous nature and the high temperature necessary to keep its solutions fluid. We have found that globular proteins such as the serum and egg albumins can be cross-linked so as to yield blocks which are suitable for ultrathin sectioning.


Author(s):  
H. Koike ◽  
S. Sakurai ◽  
K. Ueno ◽  
M. Watanabe

In recent years, there has been increasing demand for higher voltage SEMs, in the field of surface observation, especially that of magnetic domains, dislocations, and electron channeling patterns by backscattered electron microscopy. On the other hand, the resolution of the CTEM has now reached 1 ∼ 2Å, and several reports have recently been made on the observation of atom images, indicating that the ultimate goal of morphological observation has beem nearly achieved.


Author(s):  
Mircea Fotino

A new 1-MeV transmission electron microscope (Model JEM-1000) was installed at the Department of Molecular, Cellular and Developmental Biology of the University of Colorado in Boulder during the summer and fall of 1972 under the sponsorship of the Division of Research Resources of the National Institutes of Health. The installation was completed in October, 1972. It is installed primarily for the study of biological materials without many of the limitations hitherto unavoidable in standard transmission electron microscopy. Only the technical characteristics of the installation are briefly reviewed here. A more detailed discussion of the experimental program under way is being published elsewhere.


Author(s):  
Leon Dmochowski

Electron microscopy has proved to be an invaluable discipline in studies on the relationship of viruses to the origin of leukemia, sarcoma, and other types of tumors in animals and man. The successful cell-free transmission of leukemia and sarcoma in mice, rats, hamsters, and cats, interpreted as due to a virus or viruses, was proved to be due to a virus on the basis of electron microscope studies. These studies demonstrated that all the types of neoplasia in animals of the species examined are produced by a virus of certain characteristic morphological properties similar, if not identical, in the mode of development in all types of neoplasia in animals, as shown in Fig. 1.


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