Purification of human immune interferon
AssigneeMeloy Laboratories, Inc.
InventorIrwin A. Braude
Life of this patent
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Abstract
Disclosed is a process for the purification of crude immune interferon to a near homogeneous preparation which comprises: (a) adsorbing the crude interferon onto a column containing Controlled Pore Glass beads and eluting with ammonium sulfate, (b) adsorbing the interferon containing eluant onto a column containing either Concanavalin A-Sepharose, lentil lectin-Sepharose or pea lectin-agarose and eluting with a buffer containing a sugar, (c) adsorbing the interferon containing eluant onto a column containing Heparin-Sepharose or Procian Red-agarose and eluting with a high salt content buffer, and (d) treating the interferon containing eluant in a gel-filtration column equilibrated in high salt to obtain a solution of immune interferon that is nearly homogeneous.
Claims17
A process for purifying crude human immune interferon which comprises:
- (a) contacting a solution of crude interferon with Controlled Pore Glass beads for a period of time sufficient to accomplish equilibrated adsorption of the crude interferon to the beads in the presence of a neutral pH buffer, washing the glass beads having interferon adsorbed thereon with a chemically compatable buffer to remove the unbound contaminant until the optical density of the eluate at 280 nm is about 0, washing the glass beads having interferon adsorbed thereon with a neutral pH buffer solution until the optical density at 280 nm is about 0, eluting the adsorbed immune interferon from the glass beads with an ammonium sulfate solution;
- (b) contacting the solution of interferon with an absorbent selected from the group consisting of Concanavalin A-Sepharose, lentil lectin-Sepharose and pea lectin-agarose for the period of time sufficient to accomplish equilibrated adsorption of the interferon to the absorbent in the presence of a neutral pH buffer, washing the absorbent having interferon adsorbed thereon from the soluble fraction with ammonium sulfate until the optical density of the eluate at 280 nm is about 0, washing the absorbent with a neutral pH buffer until the optical density of the eluate at 280 nm is about 0, eluting the adsorbed interferon with a buffer containing a member of the group consisting of alpha-methyl D-mannoside and 1-methyl D-glucoside;
- (c) contacting the interferon with a member selected from the group consisting of Heparin-Sepharose or Procian Red-agarose for a period of time sufficient to accomplish equilibrated adsorption of the interferon to the member equilibrated in a neutral pH buffered solution, washing the member containing adsorbed interferon thereon with first a neutral pH buffered solution containing a sugar selected from the group consisting of alpha-methyl D-mannoside and 1-methyl D-glucoside to remove the unbound material until the optical density of the eluate at 280 nm is about 0, washing with a neutral pH buffered solution until the optical density is about 0 to remove the bound material eluting the adsorbed interferon with a second neutral pH buffered solution containing a highly concentrated salt, and;
- (d) further purifying by subjecting the elution fraction to gel-filtration treatment equilibrated in a neutral pH buffered solution containing a highly concentrated salt, and eluting whereby the solutes within the elution fractions are separated according to their molecular weights;
- and collecting the solute fraction containing a near homogeneous interferon preparation.
A process for purifying crude human immune interferon which comprises, contacting a solution of crude interferon with a resin from the group consisting of Heparin-Sepharose or Procian-Red agarose for a period of time sufficient to accomplish equilibrated adsorption of the interferon to the resin, washing the resin interferon adsorbed thereon with phosphate buffered saline containing sugar selected from the group consisting of alpha-methyl D-mannoside, and 1-methyl D-glucoside until the optical density at 280 nm is about 0, washing with phosphate buffered saline solution until the optical density at 280 nm is about 0, and eluting the adsorbed interferon with phosphate buffered saline containing 2 M sodium chloride.
In a process for purifying human immune interferon comprising contacting a solution of crude interferon with Controlled Pore Glass beads for a period of time sufficient to accomplish equilibrated adsorption of the crude interferon to the beads;
- separating the adsorbed interferon from the glass beads, the improvement comprising;
- eluting the adsorbed human immune interferon from the Controlled Pore Glass beads with a solution containing ammonium sulfate.
Dependencies are inferred from the claim text.