scholarly journals Development of a Lyophilization Process for Campylobacter Bacteriophage Storage and Transport

2020 ◽  
Vol 8 (2) ◽  
pp. 282 ◽  
Author(s):  
Lu Liang ◽  
Nicholas B. Carrigy ◽  
Samuel Kariuki ◽  
Peter Muturi ◽  
Robert Onsare ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Relative Humidity ◽  
Pathogenic Bacteria ◽  
Drying Process ◽  
Transport Costs ◽  
Food And Agriculture ◽  
Transmission Electron ◽  
Freeze Dried ◽  
International Transport

Bacteriophages are a sustainable alternative to control pathogenic bacteria in the post-antibiotic era. Despite promising reports, there are still obstacles to phage use, notably titer stability and transport-associated expenses for applications in food and agriculture. In this study, we have developed a lyophilization approach to maintain phage titers, ensure efficacy and reduce transport costs of Campylobacter bacteriophages. Lyophilization methods were adopted with various excipients to enhance stabilization in combination with packaging options for international transport. Lyophilization of Eucampyvirinae CP30A using tryptone formed a cake that limited processing titer reduction to 0.35 ± 0.09 log10 PFU mL−1. Transmission electron microscopy revealed the initial titer reduction was associated with capsid collapse of a subpopulation. Freeze-dried phages were generally stable under refrigerated vacuum conditions and showed no significant titer changes over 3 months incubation at 4 °C (p = 0.29). Reduced stability was observed for lyophilized phages that were incubated either at 30 °C under vacuum or at 4 °C at 70% or 90% relative humidity. Refrigerated international transport and rehydration of the cake resulted in a total phage titer reduction of 0.81 ± 0.44 log10 PFU mL−1. A significantly higher titer loss was observed for phages that were not refrigerated during transport (2.03 ± 0.32 log10 PFU mL−1). We propose that lyophilization offers a convenient method to preserve and transport Campylobacter phages, with minimal titer reduction after the drying process.

scholarly journals Correlation between structure and mass distribution of the nuclear pore complex and of distinct pore complex components.

1990 ◽  
Vol 110 (4) ◽  
pp. 883-894 ◽  
Author(s):  
R Reichelt ◽  
A Holzenburg ◽  
E L Buhle ◽  
M Jarnik ◽  
A Engel ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Nuclear Envelope ◽  
Three Dimensional ◽  
Structural Features ◽  
Nuclear Pore ◽  
Xenopus Laevis Oocyte ◽  
Three Dimensional Model ◽  
Transmission Electron ◽  
Freeze Dried

Nuclear pore complexes (NPCs) prepared from Xenopus laevis oocyte nuclear envelopes were studied in "intact" form (i.e., unexposed to detergent) and after detergent treatment by a combination of conventional transmission electron microscopy (CTEM) and quantitative scanning transmission electron microscopy (STEM). In correlation-averaged CTEM pictures of negatively stained intact NPCs and of distinct NPC components (i.e., "rings," "spoke" complexes, and "plug-spoke" complexes), several fine structural features arranged with octagonal symmetry about a central axis could reproducibly be identified. STEM micrographs of unstained/freeze-dried intact NPCs as well as of their components yielded comparable but less distinct features. Mass determination by STEM revealed the following molecular masses: intact NPC with plug, 124 +/- 11 MD; intact NPC without plug, 112 +/- 11 MD; heavy ring, 32 +/- 5 MD; light ring, 21 +/- 4 MD; plug-spoke complex, 66 +/- 8 MD; and spoke complex, 52 +/- 3 MD. Based on these combined CTEM and STEM data, a three-dimensional model of the NPC exhibiting eightfold centrosymmetry about an axis perpendicular to the plane of the nuclear envelope but asymmetric along this axis is proposed. This structural polarity of the NPC across the nuclear envelope is in accord with its well-documented functional polarity facilitating mediated nucleocytoplasmic exchange of molecules and particles.

Localization of Silver in the Spleen of Argyric Rats by Energy Dispersive X-Ray Analysis Coupled with Scanning and Transmission Electron Microscopy

1977 ◽  
Vol 35 ◽  
pp. 504-506
Author(s):  
G. Pereira
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Computer Assisted ◽  
White Pulp ◽  
Reticular Cell ◽  
X Ray ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Electron

Previous electron microscopic observations of the spleen have revealed the white pulp to be completely separated from the extravasated blood in the surrounding marginal zone by a strategically-located, double layer of reticular cells ensheathing a coarse reticular fiber. Similarly, a single reticular cell layer has been observed to form a continuous investment for all white pulp capillaries. To test the significance of this apparent isolation of the splenic white pulp from the blood, the distribution and composition of silver deposits in the spleen of argyric rats were determined by transmission and scanning electron microscopy coupled with computer-assisted x-ray analysis.Young male albino rats were made argyric by supplying them for many months with drinking water to which 1.5gm per liter of silver nitrate had been added. Specimens from the spleens of control and argyric animals were prepared for conventional transmission electron microscopy by glutaraldehyde-osmium fixation. For scanning electron microscopy, other specimens were fixed in buffered glutaraldehyde, freeze-dried in vacuo, coated with a thin film of gold- palladium and examined in a Cambridge Stereoscan Mark II.

scholarly journals Solution and surface effects on plasma fibronectin structure.

1983 ◽  
Vol 97 (6) ◽  
pp. 1686-1692 ◽  
Author(s):  
N M Tooney ◽  
M W Mosesson ◽  
D L Amrani ◽  
J F Hainfeld ◽  
J S Wall
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Ionic Strength ◽  
Scanning Transmission Electron Microscopy ◽  
Plasma Fibronectin ◽  
Scanning Transmission Electron ◽  
Neutral Ph ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission

As assessed by electron microscopy, the reported shape of the plasma fibronectin molecule ranges from that of a compact particle to an elongated, rod-like structure. In this study, we evaluated the effects of solution and surface conditions on fibronectin shape. Freeze-dried, unstained human plasma fibronectin molecules deposited at pH 7.0-7.4 onto carbon films and examined by scanning transmission electron microscopy appeared relatively compact and pleiomorphic, with approximate average dimensions of 24 nm X 16 nm. Negatively stained molecules also had a similar shape but revealed greater detail in that we observed irregular, yarn-like structures. Glutaraldehyde-induced intramolecular cross-linking did not alter the appearance of plasma fibronectin. Molecules deposited at pH 2.8, pH 9.3, or after succinylation were less compact than those deposited at neutral pH. In contrast, fibronectin molecules sprayed onto mica surfaces at pH 7, rotary shadowed, and examined by transmission electron microscopy were elongated and nodular with a contour length of 120-130 nm. Sedimentation velocity experiments and electron microscopic observations indicate that fibronectin unfolds when it is succinylated, when the ionic strength is raised at pH 7, or when the pH is adjusted to 9.3 or 2.8. Greater unfolding is observed at pH 2.8 at low ionic strength (less than 0.01) compared with material at that pH in 0.15 M NaCl solution. We conclude that (a) the shape assumed by the fibronectin molecule can be strongly affected by solution conditions and by deposition onto certain surfaces; and that (b) the images of fibronectin seen by scanning transmission electron microscopy at neutral pH on carbon film are representative of molecules in physiologic solution.

Analysis of Molluscan Hemocyanins by Scanning Transmission Electron Microscopy and Light Scattering

1988 ◽  
Vol 46 ◽  
pp. 162-163
Author(s):  
P. S. Furcinitti ◽  
J. S. Wall ◽  
M. G. Hamilton ◽  
T. T. Herskovits
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Light Scattering ◽  
Scanning Transmission Electron Microscopy ◽  
Large Angle ◽  
Scattered Electron ◽  
Scanning Transmission Electron ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission

Hemocyanins are copper containing, oxygen binding proteins found in many invertebrate species of the phyla Arthropoda and Mollusca. Molluscan hemocyanins are cylindrical macromolecular assemblies of a basic decameric unit. The hemocyanins of two chitons, Stenonlax conspicua and Mopalia muscosa. are decamers, while those of two gastropods, Fasciolaria tulipa and Pleuroplora gieantea. are di-decamers. The hemocyanins of a third gastropod species, Busvcon contrarium can exist in a spectrum of multi-decameric forms. Molecular weights of the various types of molluscan hemocyanins were measured by absolute light scattering and Scanning Transmission Electron Microscopy (STEM) as a first step in understanding multi-decamer formation in some types of hemocyanins.Hemocyanins were prepared as previously described, dialyzed into 1M HEPES buffer, pH 7.4, containing 0.05M MgCl and freeze-dried or negatively stained for STEM analysis. Specimens were examined at the Brookhaven STEM Biotechnology Resource, which was operated at 40 kV using a -140°C cold stage. The elastically scattered electron signal from the STEM large angle annular detector was used to form the images.

scholarly journals Studies on the structure of bovine factor V by scanning transmission electron microscopy

Blood ◽  
1985 ◽  
Vol 65 (5) ◽  
pp. 1158-1162 ◽  
Author(s):  
MW Mosesson ◽  
ME Nesheim ◽  
J DiOrio ◽  
JF Hainfeld ◽  
JS Wall ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Scanning Transmission Electron Microscopy ◽  
Flexible Structures ◽  
Factor V ◽  
Scanning Transmission Electron ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission ◽  
Bovine Factor

Abstract We studied purified bovine factor V (mol wt 330,000) by scanning transmission electron microscopy (STEM) of freeze-dried unstained or negatively contrasted preparations. Freeze-dried molecules revealed discrete shapes ranging from roughly spheroidal (100 to 120 nm) to oblong (140 to 200 nm in length X 50 to 100 nm in width). Oblong shapes could often be resolved into two or three distinct domains, ranging from 60 to 100 nm in diameter. A “satellite” nodular structure (30 to 50 nm in diameter) connected to the main molecule by a thin stalk (approximately 10 nm wide) up to 80 nm in length was occasionally seen. Glutaraldehyde-treated preparations yielded the same shapes as were seen in unfixed preparations but revealed better definition of submolecular features and “satellite” nodules. STEM mass analysis confirmed that each of the different shapes represented a monomolecular form of factor V. Negatively stained images revealed objects having the same general shapes as freeze-dried molecules, although greater detail was evident. Some images suggested that molecules consist of five or more discrete parts. Taken together, these observations indicate that factor V molecules are multidomainal, flexible structures that tend to have an irregular oblong shape with an axial ratio between 3:2 and 2:1.

scholarly journals Studies on the structure of bovine factor V by scanning transmission electron microscopy

Blood ◽  
1985 ◽  
Vol 65 (5) ◽  
pp. 1158-1162
Author(s):  
MW Mosesson ◽  
ME Nesheim ◽  
J DiOrio ◽  
JF Hainfeld ◽  
JS Wall ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Scanning Transmission Electron Microscopy ◽  
Flexible Structures ◽  
Factor V ◽  
Scanning Transmission Electron ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission ◽  
Bovine Factor

We studied purified bovine factor V (mol wt 330,000) by scanning transmission electron microscopy (STEM) of freeze-dried unstained or negatively contrasted preparations. Freeze-dried molecules revealed discrete shapes ranging from roughly spheroidal (100 to 120 nm) to oblong (140 to 200 nm in length X 50 to 100 nm in width). Oblong shapes could often be resolved into two or three distinct domains, ranging from 60 to 100 nm in diameter. A “satellite” nodular structure (30 to 50 nm in diameter) connected to the main molecule by a thin stalk (approximately 10 nm wide) up to 80 nm in length was occasionally seen. Glutaraldehyde-treated preparations yielded the same shapes as were seen in unfixed preparations but revealed better definition of submolecular features and “satellite” nodules. STEM mass analysis confirmed that each of the different shapes represented a monomolecular form of factor V. Negatively stained images revealed objects having the same general shapes as freeze-dried molecules, although greater detail was evident. Some images suggested that molecules consist of five or more discrete parts. Taken together, these observations indicate that factor V molecules are multidomainal, flexible structures that tend to have an irregular oblong shape with an axial ratio between 3:2 and 2:1.

Cuticular Pores in a Marine Mite

1972 ◽  
Vol 30 ◽  
pp. 164-165
Author(s):  
John H. Crowe
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Fine Structure ◽  
Epoxy Resin ◽  
Low Viscosity ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Electron

Several species of marine mites are known to possess elaborate cuticular pores, the function of which is not known (Newell, 1947). In the present investigation the fine structure of the cuticular pores in a Halacarid mite, Copidognathus curtusi was studied, with a view towards discovering the function of the pores.For scanning electron microscopy the animals were fixed in cacodylatebuffered glutaraldehyde, post-fixed in osmium, and freeze-dried. For transmission electron microscopy they were fixed as above, embedded in Spurr's low viscosity epoxy resin, and sectioned on diamond knives.C. curtusi was chosen for study because of the abundance of pores in its cuticle.

Microscopy and microanalysis of calcification in the developing and adult barnacle

1995 ◽  
Vol 53 ◽  
pp. 700-701
Author(s):  
Crystal M. Brimer ◽  
Edward Roberts ◽  
Daniel Hockett ◽  
Craig C. Freudenrich ◽  
Ann LeFurgey
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Time Course ◽  
Shell Growth ◽  
Cellular Level ◽  
Transmission Electron ◽  
Freeze Dried ◽  
Scanning Transmission ◽  
Juvenile Stages ◽  
Electron Imaging

The barnacle begins its development in its first larval stage, the naupilus, and matures to the larval cyprid stage. Permanent attachment of the barnacle occurs between the cyprid and juvenile stages. Soon after, calcification takes place and the shells of the barnacle grow upward. The application of secondary electron imaging (SEI) and electron probe microanalysis (EPXMA) has defined the time course of calcification and shell growth in the barnacle. Freeze-dried cryosections of the larval cyprid stage can also be employed for localization of calcium at the cellular level with scanning transmission electron microscopy (STEM)/EPXMA; however these sections are unstained, thereby offering very little contrast and making the recognition of ultrastuctural compartments difficult. In an attempt to identify these structures, conventional fixation staining and transmission electron microscopy (TEM) will be used. The light microscopic study of the cyprid has been extensive, but very little ultrastructural research has been conducted. The goal of this study is twofold: to use light microscopy (LM) and TEM as guides in identifying structures of the STEM/EPXMA cryosections; and to follow the growth of the barnacle by using STEM/EPXMA to detect calcium tracers in the shell.

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