Comparison of different ligand densities for the manufacture of CB Hep-1 immunosorbents.
Different ligand densities of monoclonal antibody (Mab) CB.Hep-1 were studied during covalent coupling on Sepharose CL-4B for recombinant hepatitis B surface antigen (rHBsAg) immunoaffinity purification. Ligand densities of 2.2, 3.2, 4.2 and 5.2 mg Mab/ml immunosorbents, respectively, were assayed during five cycles of immunoaffinity chromatography (IAC). Adsorption capacities averaged either 3.2 mg/ml (0.57 mg rHBsAg/ml immunosorbent/5.42 mg of total purified protein) or 5.2 mg/ml (0.56 mg rHBsAg/ml immunosorbent/5.05 mg total purified protein). Immunosorbents showed ligand leakage levels below 3 ng Mab/microg rHBsAg. Antigen purity was higher than 95% in all cases. The results suggest that a ligand density (LD) of 3.2 mg Mab/ml immunosorbent should be used for immunoaffinity chromatography because no significant differences were found in the ligand densities studied (P-value=0.012), which saves 40% of CB.Hep-1 immunosorbent manufacturing cost in comparison with 5 mg Mab/ml immunosorbent, which is currently used in large-scale production.
Gómez
L
L
Monoclonal Antibodies Department, Center for Genetic Engineering and Biotechnology, Ave 31/158 and 190, P O Box 6162, Havana 10600, Cuba. leonardo.gomez@cigb.edu.cu
Hernández
R
R
Ibarra
N
N
Valdés
R
R
Campos
Y
Y
Tamayo
A
A
Fernández
D
D
Figueroa
A
A
Alvarez
T
T
Montero
J A
JA
D000911:Antibodies, Monoclonal / Q000032:analysis / Q000096:biosynthesis / Q000737:chemistry* / Q000276:immunology; D000936:Antigen-Antibody Complex / Q000737:chemistry / Q000276:immunology; D002846:Chromatography, Affinity / Q000379:methods*; D006514:Hepatitis B Surface Antigens / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification*; D007163:Immunosorbent Techniques; D008024:Ligands; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D006514:Hepatitis B Surface Antigens; D008024:Ligands; D011994:Recombinant Proteins