Partial Purification of Human Lymphocyte-Activating Factor (LAF) by Ultrafiltration and Electrophoretic Techniques

Lymphocyte-activating factor (LAF) has been produced by culturing human peripheral blood leukocytes in the presence of lipopolysaccharide and autologous human serum. The major LAF activity, identifiable by Sephadex chromatography (m.w. 13,000) was separated from most serum proteins in the culture me...

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Published inThe Journal of immunology (1950) Vol. 119; no. 6; pp. 2019 - 2023
Main Authors Lachman, Lawrence B, Hacker, Miles P, Handschumacher, Robert E
Format Journal Article
LanguageEnglish
Published United States Am Assoc Immnol 01.12.1977
Subjects
Animals
Electrophoresis, Polyacrylamide Gel
Female
Guinea Pigs
Isoelectric Focusing
Leukocytes - immunology
Lipopolysaccharides - pharmacology
Lymphokines - isolation & purification
Ultrafiltration
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Summary:Lymphocyte-activating factor (LAF) has been produced by culturing human peripheral blood leukocytes in the presence of lipopolysaccharide and autologous human serum. The major LAF activity, identifiable by Sephadex chromatography (m.w. 13,000) was separated from most serum proteins in the culture medium by ultrafiltration with a hollow fiber device. Sucrose gradient isoelectric focusing of the concentrated ultrafiltrate yielded a single peak of LAF activity with an average isoelectric point of pH 6.8. Semi-preparative polyacrylamide gel electrophoresis of the isoelectric focusing sample resulted in separation of the LAF activity from detectable amounts of serum proteins. The recovered LAF activity was estimated to be purified more than 16,000-fold and is judged to be active in submicrogram amounts.
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ISSN:0022-1767
1550-6606
DOI:10.4049/jimmunol.119.6.2019