G. Hunt et al., CAPILLARY ISOELECTRIC-FOCUSING AND SODIUM DODECYL-SULFATE CAPILLARY GEL-ELECTROPHORESIS OF RECOMBINANT HUMANIZED MONOCLONAL-ANTIBODY HER2, Journal of chromatography, 744(1-2), 1996, pp. 295-301
Citations number
20
Categorie Soggetti
Chemistry Analytical","Biochemical Research Methods
Capillary isoelectric focusing (cIEF) and IEF of recombinant humanized
monoclonal antibody HER2 (rhuMAbHER2) show five charged isoforms with
estimated pi values ranging from 8.6-9.1, The cIEF assay demonstrated
good precision with relative standard deviations (R.S.D,) 0.7-3.7% an
d 0.4-4.2% for intra and interassay analysis, respectively. The method
was linear for the area of the main peak over the concentration range
2-250 mu g/ml with a Pearson correlation coefficient >0.99. The limit
of detection for the main peak was determined to be 2 ppm. With both
sodium dodecyl sulfate-capillary gel electrophoresis (SDS-CGE) and SDS
-polyacrylamide gel electrophoresis, the nonreduced rhuMAbHER2 migrate
d as a single major peak with minor peaks in the aggregate and clip re
gions. After reduction, the electropherogram and the slab gel showed t
he expected heavy chain and light chain fragments with minor peaks in
the aggregate and clip regions. The SDS-CGE assay showed good precisio
n with R.S.D, values of 0.1-7.8% and 0.1-8.1% for intra and interassay
analysis, respectively, The Pearson correlation coefficient for the a
rea of the main peak was >0.99 demonstrating linearity for the concent
ration range 0.5-500 mu g/ml. The limit of detection for intact rhuMAb
HER2 was determined To be 0.5 ppm. The data presented demonstrates the
feasibility of replacing the slab gel techniques with capillary elect
rophoresis in a quality control environment.