Absorption of anti-blood group A antibodies on P-selectin glycoprotein ligand-1/immunoglobulin chimeras carrying blood group A determinants: core saccharide chain specificity of the Se and H gene encoded alpha1,2 fucosyltransferases in different host cells.
To specifically eliminate recipient anti-blood group ABO antibodies prior to ABO-incompatible organ or bone marrow transplantation, an efficient absorber of ABO antibodies has been developed in which blood group determinants may be carried at high density and by different core saccharide chains on a mucin-type protein backbone. The absorber was made by transfecting different host cells with cDNAs encoding a P-selectin glycoprotein ligand-1/mouse immunoglobulin G(2b) chimera (PSGL-1/mIgG(2b)), the H- or Se-gene encoded alpha1,2-fucosyltransferases (FUT1 or FUT2) and the blood group A gene encoded alpha1,3 N-acetylgalactosaminyltransferase (alpha1,3 GalNAcT). Western blot analysis of affinity-purified recombinant PSGL-1/mIgG(2b) revealed that different precursor chains were produced in 293T, COS-7m6, and Chinese hamster ovary (CHO)-K1 host cells coexpressing FUT1 or FUT2. FUT1 directed expression of H type 2 structures mainly, whereas FUT2 preferentially made H type 3 structures. None of the host cells expressing either FUT1 or FUT2 supported expression of H type 1 structures. Furthermore, the highest A epitope density was on PSGL-1/mIgG2(2b) made in CHO-K1 cells coexpressing FUT2 and the alpha1,3 GalNAcT. This PSGL-1/mIgG(2b) was used for absorption of anti-blood group A antibodies in human blood group O serum. At least 80 times less A trisaccharides on PSGL-1/mIgG(2b) in comparison to A trisaccharides covalently linked to macroporous glass beads were needed for the same level of antibody absorption. In conclusion, PSGL-1/mIgG(2b), if substituted with A epitopes, was shown to be an efficient absorber of anti-blood group A antibodies and a suitable model protein for studies on protein glycosylation.
Löfling
Jonas C
JC
Division of Clinical Immunology, F79, IMP1, Karolinska Institutet, Huddinge University Hospital AB, S-141 86 Stockholm, Sweden.
Hauzenberger
Elenor
E
Holgersson
Jan
J
D000017:ABO Blood-Group System / Q000276:immunology*; D000042:Absorption; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D018076:DNA, Complementary; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D005647:Fucosyltransferases / Q000235:genetics*; D006041:Goats; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D007136:Immunoglobulins / Q000235:genetics*; D051379:Mice; D019007:P-Selectin / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000276:immunology; D012680:Sensitivity and Specificity; D014162:Transfection; D000097631:Galactoside 2-alpha-L-fucosyltransferase
D000017:ABO Blood-Group System; D018076:DNA, Complementary; D000939:Epitopes; D005647:Fucosyltransferases; D007074:Immunoglobulin G; D007136:Immunoglobulins; D019007:P-Selectin; D011993:Recombinant Fusion Proteins; D000097631:Galactoside 2-alpha-L-fucosyltransferase