Two-step centrifugation method. A simplification of the density-gradient procedure for the purification of influenza virus

1973 ◽  
Vol 19 (5) ◽  
pp. 633-638 ◽  
Author(s):  
D. J. S. Arora ◽  
V. Pavilanis ◽  
P. Robert
Keyword(s):  
Influenza Virus ◽  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Sucrose Concentration ◽  
Gradient Centrifugation ◽  
Sucrose Density Gradient ◽  
Virus Preparation ◽  
Virus Particles ◽  
Centrifugation Method ◽  
Degree Of Purification

In the present investigation, centrifugation of an influenza virus preparation by the sucrose density-gradient method was carried out to determine the "upper" and "lower" concentrations of sucrose at which viral activity resided. Virus was suspended in an "upper" concentration of sucrose and was centrifuged, thus resulting in a suspension containing virus free from impurities. The sucrose concentration in the suspension containing virus was then reduced to the "lower" concentration of sucrose and the suspension was recentrifuged. Virus particles sedimented, leaving impurities in the suspension. As well as achieving the same degree of purification of the virus sample as obtained by density-gradient centrifugation, this method also succeeded in concentrating influenza virus.

Uranyl Acetate and Rotary Shadowing to Increase the Contrast of Small Aggregated Virus Particles

1969 ◽  
Vol 27 ◽  
pp. 424-425
Author(s):  
Lee F. Ellis ◽  
Richard M. Van Frank ◽  
Walter J. Kleinschmidt
Keyword(s):  
Electron Microscopy ◽  
Tissue Culture ◽  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Uranyl Acetate ◽  
Interferon Inducer ◽  
Virus Particles ◽  
Staining Methods ◽  
Rotary Shadowing

The extract from Penicillum stoliniferum, known as statolon, has been purified by density gradient centrifugation. These centrifuge fractions contained virus particles that are an interferon inducer in mice or in tissue culture. Highly purified preparations of these particles are difficult to enumerate by electron microscopy because of aggregation. Therefore a study of staining methods was undertaken.

The Combining Ability of Glycoproteins IIb, IIIa and Ca2+ in EDTA-Treated Nonaggregable Platelets

1983 ◽  
Vol 50 (04) ◽  
pp. 848-851 ◽  
Author(s):  
Marjorie B Zucker ◽  
David Varon ◽  
Nicholas C Masiello ◽  
Simon Karpatkin
Keyword(s):  
Density Gradient ◽  
Combining Ability ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Electrophoretic Pattern ◽  
Sucrose Density Gradient ◽  
Irreversible Loss ◽  
Sucrose Density Gradient Centrifugation ◽  
Crossed Immunoelectrophoresis ◽  
Triton X

SummaryPlatelets deprived of calcium and incubated at 37° C for 10 min lose their ability to bind fibrinogen or aggregate with ADP when adequate concentrations of calcium are restored. Since the calcium complex of glycoproteins (GP) IIb and IIIa is the presumed receptor for fibrinogen, it seemed appropriate to examine the behavior of these glycoproteins in incubated non-aggregable platelets. No differences were noted in the electrophoretic pattern of nonaggregable EDTA-treated and aggregable control CaEDTA-treated platelets when SDS gels of Triton X- 114 fractions were stained with silver. GP IIb and IIIa were extracted from either nonaggregable EDTA-treated platelets or aggregable control platelets with calcium-Tris-Triton buffer and subjected to sucrose density gradient centrifugation or crossed immunoelectrophoresis. With both types of platelets, these glycoproteins formed a complex in the presence of calcium. If the glycoproteins were extracted with EDTA-Tris-Triton buffer, or if Triton-solubilized platelet membranes were incubated with EGTA at 37° C for 30 min, GP IIb and IIIa were unable to form a complex in the presence of calcium. We conclude that inability of extracted GP IIb and IIIa to combine in the presence of calcium is not responsible for the irreversible loss of aggregability that occurs when whole platelets are incubated with EDTA at 37° C.

scholarly journals Purification of Torpedo californica post-synaptic membranes and fractionation of their constituent proteins

1980 ◽  
Vol 185 (3) ◽  
pp. 667-677 ◽  
Author(s):  
J Elliott ◽  
S G Blanchard ◽  
W Wu ◽  
J Miller ◽  
C D Strader ◽  
...  
Keyword(s):  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Sucrose Density Gradient ◽  
Selective Extraction ◽  
Synaptic Membranes ◽  
Detergent Solution ◽  
Major Step ◽  
Sucrose Density Gradient Centrifugation ◽  
Torpedo Californica

A rapid methof for preparation of membrane fractions highly enriched in nicotinic acetylcholine receptor from Torpedo californica electroplax is described. The major step in this purification involves sucrose-density-gradient centrifugation in a reorienting rotor. Further purification of these membranes can be achieved by selective extraction of proteins by use of alkaline pH or by treatment with solutions of lithium di-idosalicylate. The alkali-treated membranes retain functional characteristics of the untreated membranes and in addition contain essentially only the four polypeptides (mol.wts. 40000, 50000, 60000 and 65000) characteristic of the receptor purified by affinity chromatography. Dissolution of the purified membranes or of the alkali-treated purified membranes in sodium cholate solution followed by sucrose-density-gradient centrifugation in the same detergent solution yields solubilized receptor preparations comparable with the most highly purified protein obtained by affinity-chromatographic procedures.

Über die Spaltung physikalisch intakter Poliovirus-Teilchen in Nucleinsäure und leere Proteinhüllen durch Wärmebehandlung

1965 ◽  
Vol 20 (9) ◽  
pp. 870-878 ◽  
Author(s):  
O. Drees ◽  
Ch. Borna
Keyword(s):  
Protein Concentration ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Sedimentation Coefficient ◽  
Sucrose Density Gradient ◽  
Virus Protein ◽  
Analytical Ultracentrifuge ◽  
Sucrose Density Gradient Centrifugation ◽  
Virus Particles ◽  
Phosphate Buffered Saline

Purified preparations of poliovirus devoid of contaminating nucleic acid and so-called “C antigen” released their particle-bound ribonucleic acid (RNA) almost quantitatively on heating at 40°C for 48 hours in phosphate-buffered saline (pH 7.6). This process occurred without disruption of the virus protein and left, in addition to the free RNA and traces of undegraded virus particles, empty protein shells in the reaction mixture.The liberated RNA sedimented in the analytical ultracentrifuge as a very diffuse boundary with sedimentation coefficients (s20, w) in the range from about 8 S to less than 1 S. The shell material which could be isolated by means of sucrose density-gradient centrifugation proved to be homogeneous in the analytical ultracentrifuge. Its sedimentation coefficient (s20,w) extrapolated to zero protein concentration was found to be 78 S.

Influence de la cytochalasine A sur la composition de fractions subcellulaires d'hyphes en croissance du Mucor mucedo L. I. Composition du plasmalemme

1988 ◽  
Vol 34 (11) ◽  
pp. 1256-1265 ◽  
Author(s):  
Abdou Ahmed Abd El Razak El Mougith ◽  
Jean-Louis Fonvieille ◽  
Robert Dargent ◽  
Jacques Rami ◽  
Jane-Marie Touzé-Soulet
Keyword(s):  
Protein Content ◽  
Density Gradient ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Sucrose Density Gradient ◽  
Point Of View ◽  
Continuous Density ◽  
Mucor Mucedo ◽  
Phospholipid Ratio ◽  
Enzymatic Markers

The plasma membrane of young hyphae of Mucor mucedo L. growing in presence or absence of cytochalasine A was isolated by continuous density gradient centrifugation using Percoll at 10% or on discontinuous sucrose density gradient. Isolated membranes were characterized by enzymatic markers and cytochemical reactions, using electron microscopy. Lipid composition and protein content were determined. From the enzymatic point of view, the cytochalasine A induced a decrease (60%) in ATPase activity and with regard to the chemical composition of the membrane, a decrease in sterol content and in the sterol – phospholipid ratio as well as a decrease in protein content and an increase in the proportion of cysteine relative to other amino acids.

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