purification of viruses
DESCRIPTION
E. &. O. O. &. Purification of viruses. Purification. To obtain the viral particles in a single form out of cell debris i.e. removal of all impurities. Purification outline. Preparation of harvest. Methods of purification. Criteria of purity. Maintenance of purified virus. - PowerPoint PPT PresentationTRANSCRIPT
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PurificationPurification
To obtain the viral particles in a single form out of cell debris i.e. removal of all impurities.
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Purification outlinePurification outline
1.Preparation of harvest.2.Methods of purification.3.Criteria of purity.4.Maintenance of purified
virus.
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Preparation of harvest
Bacteria
Plant
Animal Insect
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Methods of purificationMethods of purification
1.Filtration.2.Centrifugation.3.Adsorption.4.Precipitation.5.Solvent extraction.6.Other methods.
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Methods of purificationMethods of purification
1.Filtration.2.Centrifugation.3.Adsorption.4.Precipitation.5.Solvent extraction.6.Other methods.
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Filtration: :
1.1. UltrafiltrationUltrafiltration 2. Gel filtration2. Gel filtration
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Ultra-filtrationUltra-filtration
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Gel filtrationGel filtration
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Methods of purificationMethods of purification
1.Filtration.2.Centrifugation.3.Adsorption.4.Precipitation.5.Solvent extraction.6.Other methods.
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centrifugation
Differential
Gradient
Isopycnic Cushion
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Methods of purificationMethods of purification
1.Filtration.2.Centrifugation.3.Adsorption.4.Precipitation.5.Solvent extraction.6.Other methods.
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Adsorption: :
1.1. Ion exchangeIon exchange 2. Heamadsroption2. Heamadsroption
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Ion exchange chromatography
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HeamadsorptionHeamadsorption
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Methods of purificationMethods of purification
1.Filtration.2.Centrifugation.3.Adsorption.4.Precipitation.5.Solvent extraction.6.Other methods.
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Precipitation: :
1. Chemical1. Chemical• AmmoniumsufateAmmoniumsufate• PEGPEG• Zinc acetaeZinc acetae
2. Physical2. Physical• Isoelectric piont
(pH change)
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Methods of purificationMethods of purification
1.Filtration.2.Centrifugation.3.Adsorption.4.Precipitation.5.Solvent extraction.6.Other methods.
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Methods of purificationMethods of purification
1.Filtration.2.Centrifugation.3.Adsorption.4.Precipitation.5.Solvent extraction.6.Other methods.
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Other methodsOther methods
• Electrophoresis.• Serological methods.• Enzymatic reactions.• HPLC
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Electrophoresis
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HPLCHPLC
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Criteria of purityCriteria of purity
Chemical purityChemical purity
Physical purityPhysical purity
Biological purityBiological purity
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Criteria of purityCriteria of purity
SpectrophotomeSpectrophotometerter
IsoradiationIsoradiationInfectivity Infectivity
serologicalserological
E.M.E.M.
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Criteria of purityCriteria of purity
SpectrophotomeSpectrophotometerter
IsoradiationIsoradiationInfectivity Infectivity
serologicalserological
E.M.E.M.
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MarkingMarking
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Criteria of purityCriteria of purity
SpectrophotomeSpectrophotometerter
IsoradiationIsoradiationInfectivity Infectivity
serologicalserological
E.M.E.M.
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Infectivity::
1. Local lesion assay1. Local lesion assay 2. Plaque assay2. Plaque assay
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y = 1.9608x
020406080
100120140160180
0 2 4 6 8 10Conc.(mg/ml)
No. o
f loc
al le
sion
s
Virus preparationVirus preparation
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y = 1.9608x
020406080
100120140160180
0 2 4 6 8 10Conc.(mg/ml)
No. o
f Pla
quesث
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Criteria of purityCriteria of purity
SpectrophotomeSpectrophotometerter
IsoradiationIsoradiationInfectivity Infectivity
serologicalserological
E.M.E.M.
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Criteria of purityCriteria of purity
SpectrophotomeSpectrophotometerter
IsoradiationIsoradiationInfectivity Infectivity
serologicalserological
E.M.E.M.
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44 - -MaintenanceMaintenance::
Lyophlization
The half life of infectivity can be measured in seconds at The half life of infectivity can be measured in seconds at 60 60 ooC; minutes at 37 C; minutes at 37 ooC, hours at 20C, hours at 20ooC , days at 4 C , days at 4 ooC and C and years at -70 years at -70 ooC or lower,C or lower, (Fenner (Fenner et alet al, 1987), 1987)
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