The murine monoclonal antibody NaM26-4C6 identifies a common structure on band 3 and glycophorin C.
The murine monoclonal antibody NaM26-4C6 (IgM class), obtained from the splenocytes of a BALB/c mouse immunized with human umbilical cord red blood cells, was characterized by agglutination test and immunoblotting analysis. The structure of the NaM26-4C6 epitope was further elucidated by using a series of peptides synthesized on pins. The antibody agglutinated untreated and chymotrypsin-treated but not trypsin- or neuraminidase-treated human erythrocytes. Agglutination-inhibition test demonstrated that the antibody recognizes an epitope located on the N-terminal trypsin-sensitive portion of glycophorin C. The antibody bound on immunoblots to glycophorin C, and also to the band 3 protein and its 69-kDa N-terminal fragment but did not bind to desialylated and de-O-glycosylated glycophorin C. Peptide mapping allowed localization of the binding site on the 23-kDa N-terminal intracellular peptide of band 3. The antibody binds to the amino-acid sequences 22EDPDIP27 of band 3 protein and 15SLEPDPGM22 of glycophorin C, and residues D and P were found to be essential. The new epitope identified by NaM26-4C6 corresponds to a linear amino acid sequence located on the N-terminal intracellular portion of band 3 and to a more complex structure involving oligosaccharide chains on the N-terminal extracellular domain of GPC.
Loirat
M J
MJ
Atlantique/Vendée, Nantes, France.
Czerwinski
M
M
Duk
M
M
Blanchard
D
D
D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000276:immunology*; D000911:Antibodies, Monoclonal; D018604:Epitope Mapping; D004912:Erythrocytes / Q000276:immunology*; D006021:Glycophorins / Q000276:immunology*; D006386:Hemagglutination Tests; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D051379:Mice; D008807:Mice, Inbred BALB C; D017434:Protein Structure, Tertiary; D014470:Umbilical Cord / Q000166:cytology / Q000276:immunology
D001457:Anion Exchange Protein 1, Erythrocyte; D000911:Antibodies, Monoclonal; D006021:Glycophorins