Purification of an acid-stable bovine leukocyte interferon

• Published in: Veterinary immunology and immunopathology Vol. 29; no. 1; pp. 171 - 181
• Main Authors: Abolhassani, Mohsen, Jacobsen, Karen L.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.08.1991; Elsevier
• Subjects: Analysis of the immune response. Humoral and cellular immunity; Animals; Biological and medical sciences; BOVINAE; Cattle - blood; Chromatography, Gel; Electrophoresis, Polyacrylamide Gel; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Immunobiology; INTERFERON; Interferon Inducers; Interferon-alpha - chemistry; Interferon-alpha - isolation & purification; LEUCOCITOS; LEUCOCYTE; LEUKOCYTES; Leukocytes - metabolism; Microbiology; Miscellaneous; Molecular Weight; Regulatory factors and their cellular receptors; VIROSE; VIROSES; VIROSIS; Cell culture; Purification; Bovine; Gel electrophoresis; Induction; Leukocyte; Sendai virus; Cytokine; Biosynthesis; Paramyxoviridae; Virus; Vertebrata; Mammalia; Gel filtration; Interferon; Artiodactyla; Paramyxovirus; Ungulata
• ISSN: 0165-2427; 1873-2534
• DOI: 10.1016/0165-2427(91)90062-H

Bovine leukocyte interferon (BoL-IFN), produced in bovine peripheral blood leukocytes after priming and induction with Sendai virus, was concentrated by precipitation with KSCN (pH 3.5) and purified by gel column chromatography. Recovery of BoL-IFN from precipitation was higher when crude BoL-IFN containing more fetal bovine serum (FBS) was used. However, purity of BoL-IFN recovered from the gel filtration column was highest when crude BoL-IFN with no FBS was used. The use of 25% ethylene glycol in the column elution buffer resulted in over 93% recovery of the applied IFN activity, versus only 25% when buffer contained no ethylene glycol. Column-purified BoL-IFN was further concentrated by ultrafiltration and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in denaturing buffer. When crude BoL-IFN containing no FBS was used for purification, BoL-IFN from a selected column fraction applied to SDS-PAGE resulted in a single narrow band with an apparent molecular weight (MW) of 19 000 Da. Extraction of the SDS-PAGE gel resulted in a single peak of IFN activity indicating identity of the activity and the polypeptide. This proved to be a practical method for obtaining sufficient quantities of purified natural BoL-IFN for use in the production of monoclonal antibodies to BoL-IFN and other biological experiments.