Method of purification of viral vectors
AssigneeCanji, Inc.
InventorsPaul W. Shabram, Bernard G. Huyghe, Xiaodong Liu, H. Michael Shepard
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Abstract
The invention provides a method for purifying viral vectors containing therapeutic genes for use in gene therapy. The invention comprises a method of purification from a cell lysate of a recombinant viral vector containing a therapeutic gene, which comprises: a) treating said lysate with an enzymatic agent that selectively degrades both unencapsulated DNA and RNA; b) chromatographing the treated lysate from step a) on a first resin; and c) chromatographing the eluant from step b) on a second resin; wherein one resin is an anion exchange resin and the other is an immobilized metal ion chromatography (IMAC) resin.
Claims27
A method for purification of an intact viral particle from a cell lysate, the method comprising:
- a) treating said cell lysate which contains said intact viral particle with an enzymatic agent that selectively degrades both unencapsulated DNA and RNA;
- b) chromatographing the treated lysate from step a) on a first resin;
- and
- c) chromatographing the eluant from step b) on a second resin;
- wherein one resin is an anion exchange resin and the other is an immobilized metal ion affinity resin.
A method for purification of intact viral particles from a cell lysate, the method comprising the steps of:
- a) treating said cell lysate which contains said intact viral particle with an enzymatic agent that selectively degrades both unencapsulated DNA and RNA;
- b) chromatographing the treated lysate from step a) on a first resin, and
- c) chromatographing the eluant from step b) on a second resin;
- wherein:
- one resin is an anion exchange resin and the other is a hydrophobic interaction chromatography resin;
- or
- one resin is a cation exchange resin and the other is either a hydrophobic interaction chromatography resin or an immobilized metal ion affinity resin.
A method of determining the number of intact viral particles in a sample, the method comprising:
- a) chromatographing the sample containing the intact viral particles on an anion exchange resin;
- b) monitoring the absorbance of the eluate from the chromatography of step (a) at a selected wavelength;
- and
- c) determining the total number of intact viral particles in the sample by comparing the absorbance value obtained in step (b) to a standard curve which relates absorbance to number of viral particles.
Dependencies are inferred from the claim text.