Affinity chromatography with immunochemical detection applied to the analysis of human methionyl granulocyte colony stimulating factor in serum.
An on-line, automated, HPLC method was developed for the separation and detection of recombinant human methionyl granulocyte colony stimulating factor (GCSF) and GCSF modified with poly(ethylene glycol) (PEG-GCSF) in rat serum. The automated method consists of an initial immunoaffinity chromatography step, a microbore reverse phase separation, and postcolumn immunochemical detection. Calibration curves were constructed with a lower limit of 1.5 ng of GCSF (80 fmol) and 7.5 ng of PEG-GCSF in 120 microL of rat serum. In vivo serum samples from rats dosed with PEG-GCSF were also analyzed. The development of the method is discussed as well as its general application to the detection of metabolites of recombinant proteins in body fluids.
Miller
K J
KJ
Amgen, Inc., Thousand Oaks, California 91320-1789, USA.
Herman
A C
AC
D000818:Animals; D002138:Calibration; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D000069585:Filgrastim; D016179:Granulocyte Colony-Stimulating Factor / Q000097:blood*; D006801:Humans; D007120:Immunochemistry; D051381:Rats; D011994:Recombinant Proteins
D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; D000069585:Filgrastim