Isolation of interleukin 2 (IL-2) from human and mouse lymphocyte culture supernatants by batch adsorption onto silicic acid.
Described herein is a simple, efficient and inexpensive batch adsorption procedure for the isolation and partial purification of the hydrophobic T cell growth-promoting lymphokine interleukin 2 (IL-2) from crude culture supernatants (SN) of freshly isolated human lymphocytes and leukemic T cells of established lines including human Jurkat J6.2, Gibbon ape MLA-144 and mouse EL-4. In this method, IL-2 was isolated by batch adsorption onto microparticulate silicic acid (SA) by stir-mixing the SA with SN (10 mg/ml; 30 min; 37 degrees C). Thereafter, the SA was pelleted by centrifugation and washed twice with phosphate-buffered saline (PBS). The IL-2 was eluted by adding to the pelleted IL-2-binding SA 5 vols. of ethylene glycol (EG; 50%, v/v) in PBS (pH 7.2) with high salt (1.4 M NaCl). The lymphokine-rich concentrate was then dialyzed (6 kDa MWCO) against PBS to remove the EG and low molecular weight growth inhibitors. The application of the proposed procedure was further defined in experiments in which SA was successfully employed for recovering IL-2 from SN of cultures in which the medium had been supplemented with fetal calf (FCS) or human serum to achieve maximal lymphokine production. Also presented are the results of experiments defining the SA adsorption of proteins from whole sera (e.g., FCS, calf, human and horse) and the relative affinity of different purified proteins for this matrix (e.g., bovine serum albumin, human serum albumin, casein hydrolysate, bovine gamma-globulin and bovine beta-lactoglobulin). The proposed SA procedure may prove useful for isolating other hydrophobic immunoregulatory molecules, and a 2-step purification scheme is anticipated in which the SA adsorption procedure will be used as a preparative method preceding reverse phase high performance liquid chromatography and monoclonal antibody affinity chromatography.
D000328:Adult; D000818:Animals; D001798:Blood Proteins / Q000302:isolation & purification; D002417:Cattle; D002460:Cell Line; D002478:Cells, Cultured; D005026:Ethylene Glycols; D006801:Humans; D006922:Hylobates; D007163:Immunosorbent Techniques; D007376:Interleukin-2 / Q000302:isolation & purification* / Q000378:metabolism; D007942:Leukemia, Experimental / Q000276:immunology; D051379:Mice; D012824:Silicic Acid; D012822:Silicon Dioxide; D013601:T-Lymphocytes / Q000378:metabolism*
D001798:Blood Proteins; D005026:Ethylene Glycols; D007376:Interleukin-2; D012824:Silicic Acid; D012822:Silicon Dioxide