Fluidized-bed receptor-affinity chromatography.
A multipurpose fluidized-bed receptor-affinity purification system based upon the biological recognition between an immobilized receptor and its soluble protein ligands is described. The fluidized affinity sorbent consists of a soluble form of interleukin-2 receptor chemically bonded to an aldehyde derivative of controlled pore glass beads, which have a pore diameter of 1000 A and a particle density of 1.2-1.3 g/mL. The fluidized-bed separation device used in this study consists of a specially designed column fitted at the inlet end with a perforated distributor plate covered with a screen and the top outlet with an adjustable piston. The fluidized-bed consisting of a loose gel matrix permits the unimpeded passage of cell debris and particulate matter, while the target protein is captured by the affinity beads. Purification of the humanized-anti-Tac monoclonal antibody is used as a model system to determine the operational parameters. Also, fluidized-bed receptor-affinity chromatography has been successfully employed in the purification of recombinant interleukin-2 and single chain anti-Tac(Fv)-Pseudomonas exotoxin immunotoxin from unclarified inclusion body extracts. Overall, fluidized-bed receptor-affinity chromatography is found to be a productive affinity method suitable for the purification of recombinant human interleukin-2 and related molecules.
Roche Research Center, Hoffmann-La Roche Inc., Nutley, NJ 07110.
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C
Spence
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Schaffer
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S
Kessler
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Bailon
D036002:ADP Ribose Transferases; D000911:Antibodies, Monoclonal / Q000276:immunology; D001427:Bacterial Toxins; D002460:Cell Line; D002846:Chromatography, Affinity / Q000379:methods*; D005098:Exotoxins / Q000276:immunology; D006801:Humans; D000922:Immunotoxins / Q000302:isolation & purification; D007376:Interleukin-2 / Q000302:isolation & purification* / Q000378:metabolism; D015375:Receptors, Interleukin-2 / Q000276:immunology / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D000911:Antibodies, Monoclonal; D001427:Bacterial Toxins; D005098:Exotoxins; D000922:Immunotoxins; D007376:Interleukin-2; D015375:Receptors, Interleukin-2; D011994:Recombinant Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A