Preparative electrofocusing for industrial applications.
This paper presents a case study of using a multicompartment isoelectric focusing apparatus to determine the isoelectric points and focus preparative quantities of brain derived neurotrophic factor (BDNF) and neurotrophic factor 3 (NT3). A separation of PEGylated from unPEGylated forms of granulocyte colony stimulating factor (G-CSF) is described as well. Both BDNF and NT3 have substantially higher pI values in this system than is predicted from sequenced based modeling. Although PEGylated forms of G-CSF can be separated from the unPEGylated forms, separation of protein with differing degrees of PEGylation was not achieved. The paper additionally demonstrates that this technique can be used simultaneously as an analytical and preparative tool, eliminating the need for analytical IEF gels.
Hecht
R
R
Process Development Department, Amgen, Inc., Thousand Oaks, CA 91320-1789, USA.
Tressel
T
T
Goldshteyn
V
V
Winters
D
D
Wilson
J
J
Boone
T
T
Daniels
M
M
D019208:Brain-Derived Neurotrophic Factor / Q000032:analysis; D016179:Granulocyte Colony-Stimulating Factor / Q000032:analysis; D007525:Isoelectric Focusing / Q000295:instrumentation*; D008970:Molecular Weight; D009414:Nerve Growth Factors / Q000032:analysis; D020933:Neurotrophin 3; D011092:Polyethylene Glycols
D019208:Brain-Derived Neurotrophic Factor; D009414:Nerve Growth Factors; D020933:Neurotrophin 3; D016179:Granulocyte Colony-Stimulating Factor; D011092:Polyethylene Glycols