Practical procedures for the purification of bacterial viruses.
The efficiencies of the various methods used for phage concentration have been compared. The two-phase concentration method (with polyethylene glycol and dextran sulfate) gave maximal recoveries of infectivity for coliphages of the T-even and T-odd series and for ribonucleic acid phages and single-stranded deoxyribonucleic acid phages. Precipitation of phages by acid gave high yields when applied to T2 and T4 phages but not with T3 and T7 coliphages. Differential centrifugation was efficient when sedimented phages were gently dispersed before repeating the centrifugation cycle. The efficiencies of the various methods have also been confirmed by electron microscope studies, which also show that the two-phase concentration method gave rise to intact phages. Zone centrifugations in sucrose gradients (12.5 to 52.5%) indicated that coliphages of the T-even series sediment faster than T-odd coliphages; they may thus be separated from each other and from empty ghosts by centrifugation at 100,000 x g for 40 min. Equilibrium centrifugation in preformed cesium chloride gradients was also useful for phage concentration and purification. This study also deals with some optical properties of purified phages; optical cross sections and absorbance ratios (at 260 and 280 nm) of the various preparations are given.
D001435:Bacteriophages / Q000302:isolation & purification*; D002499:Centrifugation, Density Gradient; D002501:Centrifugation, Zonal; D002586:Cesium; D011232:Chemical Precipitation; D002712:Chlorides; D003090:Coliphages / Q000302:isolation & purification; D003911:Dextrans; D004926:Escherichia coli; D006018:Glycols; D006851:Hydrochloric Acid; D008722:Methods; D008854:Microscopy, Electron; D008982:Molybdenum; D000644:Quaternary Ammonium Compounds; D013053:Spectrophotometry; D013194:Staining and Labeling; D013395:Sucrose; D014466:Ultraviolet Rays; D014764:Viral Proteins
D002712:Chlorides; D003911:Dextrans; D006018:Glycols; D000644:Quaternary Ammonium Compounds; D014764:Viral Proteins; D002586:Cesium; D013395:Sucrose; D008982:Molybdenum; D006851:Hydrochloric Acid