Format

Send to

Choose Destination
J Chromatogr A. 2016 Apr 8;1441:44-51. doi: 10.1016/j.chroma.2016.02.068. Epub 2016 Feb 27.

Affitins as robust tailored reagents for affinity chromatography purification of antibodies and non-immunoglobulin proteins.

Author information

1
INSERM UMR 892-CRCNA, Nantes, France; CNRS UMR 6299, Nantes, France; University of Nantes, Nantes, France.
2
INSERM UMR 892-CRCNA, Nantes, France; CNRS UMR 6299, Nantes, France; University of Nantes, Nantes, France. Electronic address: frederic.pecorari@univ-nantes.fr.

Abstract

Affinity chromatography is a convenient way of purifying proteins, as a high degree of purity can be reached in one step. The use of tags has greatly contributed to the popularity of this technique. However, the addition of tags may not be desirable or possible for the production of biopharmaceuticals. There is thus a need for tailored artificial affinity ligands. We have developed the use of archaeal extremophilic proteins as scaffolds to generate affinity proteins (Affitins). Here, we explored the potential of Affitins as ligand to design affinity columns. Affitins specific for human immunoglobulin G (hIgG), bacterial PulD protein, and chicken egg lysozyme were immobilized on an agarose matrix. The columns obtained were functional and highly selective for their cognate target, even in the presence of exogenous proteins as found in cell culture media, ascites and bacterial lysates, which result in a high degree of purity (∼95%) and recovery (∼100%) in a single step. Anti-hIgG Affitin columns withstand repetitive cycles of purification and cleaning-in-place treatments with 0.25 M NaOH as well as Protein A does. High levels of Affitin productions in Escherichia coli makes it possible to produce these affinity columns at low cost. Our results validate Affitins as a new class of tailored ligands for the affinity chromatography purification of potentially any proteins of interest including biopharmaceuticals.

KEYWORDS:

Affinity chromatography; Affitin,; Antibody; Immunoglobulin; Sac7d; Sso7d

PMID:
26952369
DOI:
10.1016/j.chroma.2016.02.068
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Elsevier Science
Loading ...
Support Center