RESIN AND CHROMATOGRAPHY COLUMN THAT PURIFIES ANTIBODIES WITH PROTEASE RESISTANT SMALL PEPTIDES

Abstract

A resin for an antibody purification and a chromatography column filled with the resin are provided, where small peptides are prepared for replacing protein A and binding to antibodies as well as protein A. The small peptides are easy and inexpensive to produce and resistant to proteases. Protease-resistant antibody binding peptides are synthesized on these resins, which is configured for the antibody purification directly.

Claims

1. A resin with a peptide molecule resistant to a protease, wherein the resin is configured to bind an antibody in a liquid containing the antibody for a separation of the antibody from a molecule in the liquid.

2. The resin with the peptide molecule resistant to the protease according to claim 1, wherein after the peptide molecule resistant to the protease is synthesized on the resin, the resin is configured for an antibody purification directly without separating the peptide molecule resistant to the protease from the resin and without binding to other resins.

3. The resin with the peptide molecule resistant to the protease according to claim 1, wherein the peptide molecule resistant to the protease is synthesized completely in a D form or a form alternating a D amino acid and an L amino acid sequentially.

4. An affinity chromatography column for an antibody purification, wherein the affinity chromatography column is filled with the resin according to claim 1.

Description

BRIEF DESCRIPTION OF THE DRAWING

[0010] FIGURE: Antibody purification with protease-resistant peptide-linked resin.

DETAILED DESCRIPTION OF THE EMBODIMENTS

[0011] The 14 amino acid long peptide, which we have observed to bind to the antibody previously, was synthesized on the resin in a peptide synthesizer by solid phase peptide synthesis method. Most of the resin was separated for antibody purification, and the chemicals used in the synthesis were removed by washing with buffer solution. A small part was separated from the resin and purified by HPLC (high performance liquid chromatography). The peptide was incubated overnight at 37? C. with proteinase K. When examined by HPLC again, it was observed that the peptide protected its original structure. Resin, prepared with protease resistant antibody binding peptide, was loaded into a column. After washing with the appropriate buffer solution, Anti-TNF-alpha monoclonal antibodies were loaded. Antibodies bound to the resin were removed from the column with a solution consisting of a mixture of glycine and NaCl (In the FIGURE). After the column was washed, this process was repeated many times and it was seen that the antibodies could continue to be purified with the same efficiency. It was observed that passing the proteinase K solution through the column did not impair the antibody binding and purification efficiency of the column.

[0012] FIGURE shows antibody purification with protease-resistant peptide-linked resin. Since the amount of antibody loaded at the 2.7.sup.th minute (1) is slightly above the binding capacity of the column, it is seen that the antibodies that do not bind until approximately 20 minutes leave the column (2), and the antibodies leave the resin very quickly after the addition of elution liquid around the 43.sup.rd minute (3).