Elimination of alkaloids from plant-derived human monoclonal antibody.
A human antiviral monoclonal antibody (mAb) expressed in transgenic tobacco plants was purified from the tobacco leaf by two different methods. In one method, total protein precipitated with ammonium sulfate was applied to a Hi-Trap protein A column (column method). In the second method, leaf supernatant obtained after liquid nitrogen leaf grinding was directly immunoprecipitated using protein A-agarose beads (immunoprecipitation method). The column and immunoprecipitation methods yielded 0.52 and 0.45 microg of plant-derived mAb (mAb(P))/g, respectively, from fresh leaf tissue. The product derived using the column method exhibited higher binding activity compared to immunoprecipitation-derived product against rabies virus strain CVS-11 in ELISA. Gas chromatography/mass spectrometry analysis, which has a detection limit of 5 pg revealed no detectable levels of nicotine or other related plant alkaloids in the purified mAb(P) from either purification procedure. Thus, both purification methodologies yield mAb(P) uncontaminated with nicotine from the tobacco leaves.
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Biotechnology Foundation Laboratories at Thomas Jefferson University, 1020 Locust Street Room 346, M-85, JAH, Philadelphia, PA 19107, USA.
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Xiaochen
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Crooks
Peter A
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Koprowski
Hilary
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D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D004797:Enzyme-Linked Immunosorbent Assay; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D009538:Nicotine / Q000032:analysis*; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism*; D011233:Precipitin Tests; D011820:Rabies virus / Q000276:immunology; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D009538:Nicotine