Optimization Of Synthesis Of Antibodies Adsorbents For Blood Purification And Evaluation Of Their Properties

Protein A-based adsorbent has been widely used in treating antibody-related illness and proven effective. The paper is to synthesize Protein A-based adsorbent by using recombinant protein A with binding selectivity for antibodies. This study also investigated the synthesis parameters and the scale-up process of MEP-based adsorbents, and evaluated its properties in the removal of antibodies.Further efforts for optimal development of Protein A-based adsorbent were focused on the synthesis route, spacer, activation density and ligand coupling method. The resulting Protein A-based adsorbent could achieve a dynamic binding capacity of 8.79 mg/mL to 39.74 mg/mL of gel for antibodies in serum. This is comparable to the performance achieved by current commercial Protein A adsorbent.The performance for autoantibody removal and safety of protein A adsorbent were evaluated. The adsorbent showed poor affinity with IgG3. For the removal of autoantibody, the purpose is that protein A adsorbent could remove autoantibody effectively while normal antibodies lowly. It was found that the adsorbent with low ligand density performed good adsorption to the rheumatoid factor (RF) and anti-nuclear antibody (ANA). When the volume of serum was twice the volume of adsorbent, the adsorbent with a ligand density of 2 mg/mL could remove more than 40% of the RF. The ANA could not be detected in all 14 serum samples from different individuals after the adsorption process.In addition, the binding capacity of the absorbent for antibodies was still about 30 mg/ml of gel after 10 cycles, and non-specific adsorption of plasma components to protein A adsorbent was limited.Different amounts of AB were used could get the adsorbents with different activation density. MEP-Sepharose could achieve a dynamic binding capacity of 28.53 mg/mL of gel for antibodies in serum. The ligand density and performance of adsorbent for antibodies would not get down when the reaction was scaled up 100 times.For the removal of IgG3 and RF, MEP-Sepharose showed a better performance than protein A adsorbent. MEP-Sepharose with a ligand density of 129μmol/mL of gel could remove 89.79% of the IgG3; MEP-Sepharose with a ligand density of 78.9μmol/mL of gel could remove 90.32% of the RF. The level of IgE became negative for eleven serum samples among fifteen serum samples from different individuals after the adsorption process by MEP-Sepharose, while only three samples by protein A adsorbent.