Anti-c-myc antibody 9E10: epitope key positions and variability characterized using peptide spot synthesis on cellulose.
The 9E10 antibody epitope (EQKLISEEDL) derives from a protein sequence in the human proto-oncogen p62(c-myc) and is widely used as a protein fusion tag. This myc-tag is a powerful tool in protein localization, immunochemistry, ELISA or protein purification. Here, we characterize the myc-tag epitope by substitutional analysis and length variation using peptide spot synthesis on cellulose. The key amino acids of this interaction are the core residues LISE. The shortest peptide with a strong binding signal is KLISEEDL. Dissociation constants of selected peptide variants to the antibody 9E10 were determined. scFv constructs with the shortest possible myc-tags were successfully detected by Western blot and ELISA, giving a signal comparable to that of the original myc-tag.
Hilpert
K
K
Institut für Biochemie, Universitätsklinikum Charité, Humboldt-Universität zu Berlin, Monbijoustr. 2, 10117 Berlin, Germany.
Hansen
G
G
Wessner
H
H
Küttner
G
G
Welfle
K
K
Seifert
M
M
Höhne
W
W
D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology; D000915:Antibody Affinity; D015153:Blotting, Western; D002482:Cellulose; D018985:Epitopes, B-Lymphocyte / Q000737:chemistry* / Q000276:immunology; D007700:Kinetics; D010452:Peptide Biosynthesis; D016271:Proto-Oncogene Proteins c-myc / Q000276:immunology*; D020539:Sequence Analysis, Protein
D000911:Antibodies, Monoclonal; D018985:Epitopes, B-Lymphocyte; D016271:Proto-Oncogene Proteins c-myc; D002482:Cellulose