Purification To obtain the viral particles in a single form out of cell debris i.e. removal of all impurities.

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Presentation transcript:

Purification To obtain the viral particles in a single form out of cell debris i.e. removal of all impurities.

Purification outline 1.Preparation of harvest. 2.Methods of purification. 3.Criteria of purity. 4.Maintenance of purified virus.

Preparation of harvest Bacteria Plant Animal Insect

Methods of purification 1.Filtration. 2.Centrifugation. 3.Adsorption. 4.Precipitation. 5.Solvent extraction. 6.Other methods.

Methods of purification 1.Filtration. 2.Centrifugation. 3.Adsorption. 4.Precipitation. 5.Solvent extraction. 6.Other methods.

: Filtration : 1. Ultrafiltration 2. Gel filtration

Ultra-filtration

Gel filtration

Methods of purification 1.Filtration. 2.Centrifugation. 3.Adsorption. 4.Precipitation. 5.Solvent extraction. 6.Other methods.

centrifugation Differential Gradient Isopycnic Cushion

Methods of purification 1.Filtration. 2.Centrifugation. 3.Adsorption. 4.Precipitation. 5.Solvent extraction. 6.Other methods.

: Adsorption : 1. Ion exchange 2. Heamadsroption

Ion exchange chromatography

Heamadsorption

Methods of purification 1.Filtration. 2.Centrifugation. 3.Adsorption. 4.Precipitation. 5.Solvent extraction. 6.Other methods.

: Precipitation : 1. Chemical AmmoniumsufateAmmoniumsufate PEGPEG Zinc acetaeZinc acetae 2. Physical Isoelectric piont (pH change)

Methods of purification 1.Filtration. 2.Centrifugation. 3.Adsorption. 4.Precipitation. 5.Solvent extraction. 6.Other methods.

Methods of purification 1.Filtration. 2.Centrifugation. 3.Adsorption. 4.Precipitation. 5.Solvent extraction. 6.Other methods.

Other methods Electrophoresis. Serological methods. Enzymatic reactions. HPLC

Electrophoresis

HPLC

Criteria of purity Chemical purity Physical purity Biological purity

Criteria of purity Spectrophotometer Isoradiation Infectivity serological E.M.

Spectrophotometer Isoradiation Infectivity serological E.M.

Marking

Spectrophotometer Isoradiation Infectivity serological E.M.

: Infectivity : 1. Local lesion assay 2. Plaque assay

Virus preparation

Criteria of purity Spectrophotometer Isoradiation Infectivity serological E.M.

Spectrophotometer Isoradiation Infectivity serological E.M.

4- Maintenance: Lyophlization The half life of infectivity can be measured in seconds at 60 oC; minutes at 37 oC, hours at 20oC, days at 4 oC and years at -70 oC or lower, (Fenner et al, 1987)