Use of caprylic acid precipitation for protein purification

Abstract

In certain embodiments, the invention provides a method of purifying a protein of interest from a mixture which comprises the protein of interest and one or more contaminants, comprising: a) subjecting the mixture to a first chromatography step; b) recovering the protein of interest in an elution solution; c) adding caprylic acid to the elution solution to form a contaminant precipitate; d) removing the contaminant precipitate from the elution solution; and e) subjecting the post-precipitated elution solution to a second chromatography column, thereby purifying the protein of interest.

We claim: 
     
         1 . A method of purifying a protein of interest from a mixture which comprises the protein of interest and one or more contaminants, comprising:
 a) subjecting the mixture to a first chromatography step;   b) recovering the protein of interest in an elution solution;   c) adding caprylic acid to the elution solution to form a contaminant precipitate;   d) removing the contaminant precipitate from the elution solution; and   e) subjecting the post-precipitated elution solution to a second chromatography column, thereby purifying the protein of interest.   
     
     
         2 . The method of  claim 1 , wherein the contaminants are selected from host cell proteins, host cell metabolites, host cell constitutive proteins, nucleic acids, endotoxins, viruses, product related contaminants, lipids, media additives and media derivatives. 
     
     
         3 . The method of  claim 1 , wherein the first chromatography is selected from an affinity chromatography, an ion exchange chromatography, a hydrophobic interaction chromatography, and a mix-mode chromatography. 
     
     
         4 . The method of  claim 3 , wherein the first chromatography is a protein A affinity chromatography. 
     
     
         5 . The method of  claim 3 , wherein the first chromatography is an anion exchange chromatography. 
     
     
         6 . The method of  claim 3 , wherein the first chromatography is a cation exchange chromatography. 
     
     
         7 . The method of  claim 1 , wherein the second chromatography is selected from an ion exchange chromatography, a hydrophobic interaction chromatography, and a mix-mode chromatography. 
     
     
         8 . The method of  claim 7 , wherein the second chromatography is a cation exchange chromatography. 
     
     
         9 . The method of  claim 7 , wherein the second chromatography is an anion exchange chromatography. 
     
     
         10 . The method of  claim 1 , wherein the second chromatography is a positive-charged membrane chromatography or a hydrophobic interaction membrane chromatography. 
     
     
         11 . The method of  claim 1 , wherein the contaminant precipitate is removed by centrifugation, sterile filtration, depth filtration or tangential flow filtration. 
     
     
         12 . The method of  claim 1 , wherein the pH of the elution solution is between about 3 and 7. 
     
     
         13 . The method of  claim 18 , wherein the pH of the elution solution is between 5 and 6. 
     
     
         14 . The method of  claim 12 , wherein the pH of the elution solution is adjusted before or after the addition of caprylic acid. 
     
     
         15 . The method of  claim 1 , wherein the final concentration of the caprylic acid is at least about 0.05% (v/v). 
     
     
         16 . The method of  claim 1 , wherein the contaminant precipitate is allowed to form for at least 5 minutes after addition of the caprylic acid. 
     
     
         17 . The method of  claim 1 , wherein the mixture is not subjected to an additional chromatography step. 
     
     
         18 . The method of  claim 1 , wherein the mixture is selected from a harvested cell culture fluid, a cell culture supernatant, and a conditioned cell culture supernatant, a cell lysate, and a clarified bulk. 
     
     
         19 . The method of  claim 18 , wherein the cell culture is a mammalian cell culture. 
     
     
         20 . The method of  claim 1 , wherein the protein of interest is an antibody.