- The novel resin for purification of immunoglobulin G (Ig sepharose) by Arg biotech.
The spherical sepharose CL-6B beads were activated by epichlorohydrin in different epoxy contents and, L-histidine and imidazole as pseudo-affinity ligands were covalently immobilized to them. Some linkers with different length were synthesized for activation of sepharose and the activated sepharose beads modified with imidazole and the performance of these adsorbents in purification of immunoglobulin G from bovine milk were evaluated. Among the L-histidine bearing adsorbents, higher adsorption of IgG was obtained by adsorbent with the lower concentration of L-histidine. The highest amount of IgG adsorption was obtained by imidazole bearing adsorbent with the highest amount of imidazole and Among the adsorbents with synthesized linkers, the adsorbent with 1,2- ethanediol diglycidyl ether showed better performance and was able to purify 0.25 mg/mL IgG with high purity. All of these pseudo-affinity adsorbents were able to purify immunoglobulin G in one step process with high purity and efficiency.
Because of vital role of antibodies in the biotechnology, medicine, pharmaceutical and food industries, isolation and purification of antibodies is very important and it has increased dramatically over the past decades. Antibodies or immunoglobulins (Ig), are Y-shaped, unique and soluble glycoproteins that are secreted by lymphocyte B cells in the immune system to counteract foreign antigens. These specific antibody-antigen interactions have led to extensive applications of antibodies in various fields. Immunoglobulin G (IgG) is one of the important and interesting antibodies of bovine milk and purification of this bioactive proteins from bovine milk has attracted researcher’s attention during the last years. It is the most common group of antibodies which is the smallest antibody among the other immunoglobulins and it can be stable throughout the purification process. According to the important role of IgG for the purposes of immunoaffinity chromatography, immunotherapy, drug delivery, and treatment of immune disorders like all immunization, rheumatoid arthritis and cancer, it is very essential to isolate and purify it. A large number of methods, including precipitation, electrophoresis, filtration, ion exchange, size exclusion and affinity chromatography have been used for purification of antibodies. However, Affinity chromatography is the best method for purification of antibodies from the protein mixture. Affinity chromatography is a kind of separation technique that employs the unique and reversible interactions between a protein and related ligand to isolate the target protein. These binding properties that offered by affinity ligand are used for selective adsorption of target protein from a protein mixture. Recently for simplifying the purification process and providing high selectivity, efficiency and easy recovery of protein, considerable efforts have been made. To recompense these requirements, novel affinity methods have been developed by identifying and designing new ligands and matrixes. Protein A chromatography is the most widely used method for purification of immunoglobulins, which has some disadvantages like high cost, ligand leakage, low stability, prone to degradation and loss of antibody activity under severe elution conditions (pH=3). On the other hand, pseudo-specific affinity chromatography can be extensively used to purify biomolecules, which at the same time compensates for the defects of protein a chromatography. In pseudo-specific affinity chromatography, the interactions among the immobilized ligand with the target protein, depend on the complementarity of their shape, charge, hydrophobicity and so on. The benefits and particular properties of histidine and imidazole as pseudo-affinity ligands have made Them unique. Imidazole is present in the side chain of important molecules, including histidine, which has an important role in the structure and location of protein binding. Furthermore, they possess some specific properties such as good stability, mild hydrophobicity, and wide range of pKa values, asymmetric carbon atom and weak charge transfer due to the presence of imidazole ring.