Ribosome display selection of a murine IgG₁ Fab binding affibody molecule allowing species selective recovery of monoclonal antibodies

Sebastian Grimm, Feifan Yu, Per-Åke Nygren
Molecular Biotechnology 2011, 48 (3): 263-76
Affinity reagents recognizing constant parts of antibody molecules are invaluable tools in immunotechnology applications, including purification, immobilization, and detection of immunoglobulins. In this article, murine IgG₁, the primary isotype of monoclonal antibodies (mAbs) was used as target for selection of novel binders from a combinatorial ribosome display (RD) library of 10¹¹ affibody molecules. Four rounds of selection using three different mouse IgG₁ mAbs as alternating targets resulted in the identification of binders with broad mIgG₁ recognition and dissociation constants (K(D)) in the low nanomolar to low micromolar range. For one of the binders, denoted Z(mab25), competition in binding to full length mIgG₁ by a streptococcal protein G (SPG) fragment and selective affinity capture of mouse IgG₁ Fab fragments after papain cleavage of a full mAb suggest that an epitope functionally overlapping with the SPG-binding site in the CH₁ domain of mouse IgG₁ had been addressed. Interestingly, biosensor-based binding experiments showed that neither human IgG₁ nor bovine Ig, the latter present in fetal bovine serum (FBS) was recognized by Z(mab25). This selective binding profile towards murine IgG₁ was successfully exploited in species selective recovery of two different mouse mAbs from complex samples containing FBS, resembling a hybridoma culture supernatant.

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