Purification of recombinant insect transferrin from large volumes of cell culture medium using high capacity Ni(2+)-dipicolylamine gel.
We report the purification of secreted recombinant Manduca sexta transferrin from Spodoptera frugiperda (Sf9) cell culture medium in a single step using high capacity Ni(2+)-dipicolylamine (DPA)-Novarose gel. Although the original sample was highly diluted (approximately 10 micrograms transferrin/ml medium) and the cell culture medium contained 10% surfactant (Pluronic F68) and a lipid emulsion, we were able to recover the recombinant transferrin (1 mg protein/100 ml) under gentle elution conditions with 70% yield at > 90% homogeneity. This work demonstrates the versatility of immobilized metal ion affinity chromatography using a high metal ion capacity gel to purify a recombinant protein and illustrates the potential of this affinity technique for protein separations from large volumes of cell culture media that contain surfactants.
Winzerling
J J
JJ
Department of Biochemistry, University of Arizona, Tucson 85721, USA.
Pham
D Q
DQ
Kunz
S
S
Samaraweera
P
P
Law
J H
JH
Porath
J
J
D000818:Animals; D002846:Chromatography, Affinity / Q000379:methods*; D003470:Culture Media; D005782:Gels; D018402:Manduca / Q000737:chemistry* / Q000235:genetics; D009532:Nickel; D017924:Nucleopolyhedroviruses / Q000378:metabolism; D009942:Organometallic Compounds; D010847:Picolines; D011499:Protein Processing, Post-Translational; D011994:Recombinant Proteins / Q000302:isolation & purification*; D018411:Spodoptera; D014162:Transfection; D014168:Transferrin / Q000302:isolation & purification* / Q000378:metabolism
D003470:Culture Media; D005782:Gels; D009942:Organometallic Compounds; D010847:Picolines; D011994:Recombinant Proteins; D014168:Transferrin; C103101:nickel dipicolylamine; D009532:Nickel