Sequential antibody affinity chromatography of human leukocyte interferon

Area of Science:

• Immunology
• Biochemistry
• Protein Purification

Background:

• Crude leukocyte interferon (CIF) is a complex mixture of proteins with antiviral properties.
• Purification of biologically active interferon is crucial for therapeutic and research applications.
• Affinity chromatography is a powerful technique for isolating specific proteins.

Purpose of the Study:

• To develop an optimized affinity chromatography method for purifying crude leukocyte interferon (CIF).
• To characterize the specific activity of purified interferon.
• To investigate the relationship between antibody binding and interferon recovery.

Main Methods:

• Antibodies to CIF were immobilized on Sepharose 4B for affinity chromatography.
• Crude leukocyte interferon was chromatographed using the antibody-Sepharose column.
• Further purification involved chromatography against normal cellular proteins and concentration via affinity chromatography.

Main Results:

• A correlation was observed between the percentage of antibody binding to Sepharose 4B and interferon recovery.
• Optimal interferon recovery (50%-85% antibody binding) yielded a specific activity of 1-5 X 10(6) interferon units (IFU)/mg protein.
• A tenfold increase in 'direct' specific activity was achieved after subsequent purification steps, suggesting an 'indirect' specific activity of approximately 2 X 10(8) IFU/mg protein.

Conclusions:

• Optimized antibody affinity chromatography is effective for purifying crude leukocyte interferon.
• The purification strategy significantly enhanced the specific activity of interferon.
• The study provides insights into the purification challenges and potential specific activity of interferon.