Preparation of monoclonal antibody-ferritin conjugates of high specific activity

✍ Scribed by R. A. Rudick; E. K. Bloechl; D. W. Knutson

• Book ID: 104786839
• Publisher: Springer
• Year: 1984
• Tongue: English
• Weight: 578 KB
• Volume: 80
• Category: Article
• ISSN: 1432-119X
• DOI: 10.1007/bf00495776

✦ Synopsis

125I-monoclonal IgG anti-gamma chain antibodies were conjugated to ferritin using glutaraldehyde as a bifunctional reagent. The molar ratio of IgG:ferritin:glutaraldehyde resulting in the highest yield was determined. Free IgG was separated from IgG bound to ferritin by sucrose density gradient ultracentrifugation; free ferritin was separated from antibody-ferritin conjugates by differential salt precipitation. The IgF:ferritin molar ratio of the resulting product was 1:1.4, containing over 90% ferritin-IgG "monomers"; 70-90% of the 125I activity bound immunospecifically to sepharose-IgG or aggregated human globulin (AHG). The product was used as an immunologic EM marker for AHG. Monoclonal antibody-ferritin conjugates prepared by this method should prove useful for quantitative ultrastructural analysis of surface antigens.