Purification of a mouse monoclonal antibody against Erns protein from classical swine fever virus

Abstract

With the aim to follow large scale production of recombinant E rns from Classical Swine Fever Virus, a monoclonal specific antibody was obtained and optimized it downstream lab scale purification. The anti-E rns gamma globulins were purified through Protein A and Protein G affinity chromatography from ascites fluid. Three binding factors were assessed (pH, flow rate and buffer) to define dynamic binding capacity. The best binding condition to Protein A (6.6 mg/mL of matrix) was in phosphate buffer, pH 8.0 and linear flow rate of 78cm/h with a purity of 90 %. For Protein G, the best arrangement of factors was phosphate buffer, pH 7.0 and linear flow 178cm/h with 6.45mg/mL of adsorbed antibody with a purity of 95 %. The nProtein-A Sepharose matrix allowed 1.21 times more recovery than the nProtein-G Sepharose matrix. However, the elutions obtained with the G-protein were more pure.

Rabbit Anti-Mouse Serum Albumin Polyclonal Antibody Generation for Release Testing of Monoclonal Antibodies Produced From Murine Ascites Fluid

“…A sandwich ELISA was performed to assess the potential use of our pAb to quantify MSA traces in purified mAb samples. The method used here was similar to previous works (Fernández et al, 2019;Veitia et al, 2020;Machín et al, 2021) where authors estimated specific protein concentration. From all mAb preparations, the HepB.1 was the higher murine-contaminated (17.8 ng/mL) which correspond with the fact that this mAb product lot was declared non-conforming for quality control.…”