Affinity chromatography resins that are obtained by conjugation of matrices with proteins of bacterial origin, like protein A, are frequently used for the purification of numerous therapeutic monoclonal antibodies. This article presents the development of a biocatalytic method for the production of novel affinity resins with an immobilized mutant form of protein A via sortase A mediated reaction. The conditions for activation of the agarose Seplife 6FF matrix, selection of different types of linkers with free amino groups and conditions for immobilization of recombinant protein A on the surface of the activated matrix were studied. Finally, the basic operational properties, like dynamic binding capacity (DBC), temperature dependance of DBC and stability during the cleaning-in-place process of the affinity resin with the Gly-Gly-EDA-Gly-Gly linker, were assessed using recombinant hyperchimeric monoclonal antibodies. The main characteristics show comparable results with the widely used commercial samples.

Affinity chromatography is a widely used technique for antibody isolation. This article presents the successful synthesis of a novel affinity resin with a mutant form of protein A (BsrtA) immobilized on it as a ligand. The key aspect of the described process is the biocatalytic immobilization of the ligand onto the matrix using the sortase A enzyme. Moreover, we used a matrix with primary amino groups without modification, which greatly simplifies the synthesis process. The resulting resin shows a high dynamic binding capacity (up to 50 mg IgG per 1 mL of sorbent). It also demonstrates high tolerance to 0.1 M NaOH treatment and maintains its effectiveness even after 100 binding, elution, and sanitization cycles.

Monoclonal antibodies (mAbs) are widely used as therapeutic molecules for the treatment of serious diseases, primarily cancer. The market for mAbs is one of the fastest-growing segments of the biopharmaceuticals industry. Purification is a crucial stage in the production of mAbs. While staphylococcal protein A (SpA) affinity chromatography remains the gold standard in industrial mAb purification, its limitations—low alkaline stability and insufficient binding capacity of SpA, as well as the need for harsh acidic elution conditions—have driven extensive efforts for novel progressive affinity ligands. This review focuses on the development and performance of bacterial protein-based affinity resins for the purification of class G immunoglobulins (IgGs), including conventional proteins A and G, the promising protein L, and the more recently discovered protein M (from M. genitalium), each offering unique specificities for different antibody fragments and species. Hybrid ligands combining domains from multiple bacterial proteins are also discussed, along with next-generation synthetic alternatives such as affibodies, affimers, nanobodies, etc., as well as peptide-based or mixed-mode ligands. The key finding is that the reliable and time-tested resins like those based on protein A continue to dominate the market, although future trends also point toward smaller, more stable, and cost-effective synthetic ligands for specific applications.