N terminal sequencing for practical detection of monoclonal antibody.
- Author:
Wei GUO
;
Chuanfei YU
;
Meng LI
;
Lan WANG
;
Feng ZHANG
;
Chunyu LIU
;
Wenbo WANG
;
Kai GAO
- Publication Type:Journal Article
- MeSH:
Amino Acid Sequence;
Antibodies, Monoclonal;
isolation & purification;
Chromatography, Ion Exchange;
Humans;
Isoelectric Focusing;
Mass Spectrometry;
Peptide Mapping;
Peptides;
Sequence Analysis, Protein;
methods
- From:
Chinese Journal of Biotechnology
2014;30(9):1473-1480
- CountryChina
- Language:Chinese
-
Abstract:
Here we discuss whether N terminal sequencing is appropriate as one of the conventional control methods for monoclonal antibody products. We determined the N terminal sequences of two monoclonal antibody products targeting two antigens separately with both Edman degradation and mass peptide spectrometry. We also identified the characteristic peptide fragments with mass spectrometry. Furthermore, we analyzed their heterogeneity with ion exchange chromatography, capillary zone electrophoresis and Imaged Capillary Isoelectric Focusing. Edman degradation method showed that the N terminal 15 amino acids of heavy and light chains of the two monoclonal antibodies were identical. Peptide mass spectrometry demonstrated that T1 peptide fragments of heavy and light chains of the two antibodies were also the same. But in contrast, peptide mapping and the three analytical methods for heterogeneity analysis could effectively identify and differentiate the two antibodies. The N terminal sequences of two monoclonal antibodies are identical because the number of framework sequences of humanized or human monoclonal antibodies is relatively limited, so whether N terminal sequencing analysis could be regulated as one of the practical control methods should be carefully discussed. Our work also proves that the above analytical methods could combinatorially applied to the identification of monoclonal antibody products, and are more objective compared to N terminal sequencing.
