Purification of human interleukin 1 by high-performance liquid chromatography.
The utility of high-performance liquid chromatography (HPLC) in the purification of Interleukin 1 (IL 1, lymphocyte-activating factor) has been investigated Human IL 1-containing supernatants were concentrated by lyophilization and desalted using Bio-Gel P-6 DG desalting gel. Subsequently, the sample containing IL 1 activity was subjected to HPLC with a novel HPHT hydroxylapatite column. Using a sodium phosphate gradient, IL 1 was eluted as a single peak of activity separated from the major protein contaminant, yielding 90% recovery and a specific activity of 6.3 X 10(4) U/mg. Pooled fractions from Bio-Gel HPHT were concentrated and subjected either to Bio-Sil IEX 540 DEAE anion-exchange or Bio-Sil TSK 125 size exclusion chromatography. From DEAE the IL 1 activity was eluted before a linear sodium chloride gradient was started, whereas the protein contaminant was eluted at 110 mM NaCl. When TSK was used IL 1 activity was eluted within a molecular weight range of 20,000-10,000. Fractions from the DEAE or TSK columns that were positive for IL 1 activity did not contain detectable protein, suggesting a good resolution. Furthermore, the recovery from DEAE was 26% whereas TSK 125 yielded 119% of the original activity. The specific activities were 6 X 10(7) and 2.5 X 10(8) U/mg, respectively. Thus, this method provides a rapid and reproducible procedure for the purification if IL 1 for further biological characterization.
D002478:Cells, Cultured; D002848:Chromatography, DEAE-Cellulose / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006801:Humans; D007375:Interleukin-1 / Q000302:isolation & purification*; D008214:Lymphocytes / Q000378:metabolism; D012709:Serum Albumin / Q000032:analysis
D007375:Interleukin-1; D012709:Serum Albumin