Epitope tag for affinity-based applications

Patent metadata

Field	Value
Publication number	US-9518084-B2
Application number	US-201113698624-A
Country	US
Kind code	B2
Filing date	May 25, 2011
Priority date	May 25, 2010
Publication date	Dec 13, 2016
Grant date	Dec 13, 2016

Abstract

Described is an epitope tag useful in affinity-based applications. The invention further includes fusion proteins, methods for preparing fusion proteins, nucleic acid molecules encoding these fusion proteins and recombinant host cells that contain these nucleic acid molecules. The invention also relates to nanobodies and other affinity ligands specifically recognizing the epitope tag, and uses thereof in affinity-based applications.

First claim

The invention claimed is:

1. A method for purifying a non-naturally occurring fusion protein comprising (i) an epitope tag consisting of SEQ ID NO:1 and (ii) a target protein, the method comprising:
a) applying a solution containing the non-naturally occurring fusion protein to a solid support possessing an immobilized affinity ligand that binds to an epitope tag consisting of SEQ ID NO:1, wherein the solid support is a column or a bead, and wherein the affinity ligand is an antibody, antibody fragment, or single-domain antibody;
b) forming a complex between the immobilized affinity ligand and the non-naturally occurring fusion protein;
c) removing weakly bound or unbound molecules; and
d) eluting bound molecules.

2. The method of claim 1 , wherein the solid support is a bead.

3. A method for immobilizing and/or capturing a non-naturally occurring fusion protein on a solid support, wherein the non-naturally occurring fusion protein comprises (i) an epitope tag consisting of SEQ ID NO:1 and (ii) a target protein, the method comprising:
a) applying a solution containing a non-naturally occurring fusion protein to a solid support possessing an immobilized affinity ligand that binds to an epitope tag consisting of SEQ ID NO:1, wherein the solid support is a column or a bead, and wherein the affinity ligand is an antibody, antibody fragment, or single-domain antibody;
b) forming a complex between the immobilized affinity ligand and the fusion protein; and
c) removing weakly bound or unbound molecules.

4. The method of claim 3 , wherein the solid support is a bead.

5. The method of claim 3 , wherein the solid support is a column.

6. The method of claim 1 , wherein the solid support is a column.

Assignees

• Pardon Els
• Steyaert Jan
• Wyns Lode
• Vib Vzw
• Univ Brussel Vrije

Inventors

• Pardon Els
• Steyaert Jan
• Wyns Lode

Classifications

• C07K5/1021 Primary

  with the first amino acid being acidic · CPC title

• C07K1/22

  Affinity chromatography or related techniques based upon selective absorption processes · CPC title

• C07K2319/40

  containing a tag for immunodetection, or an epitope for immunisation · CPC title