Background: Lewis phenotyping by hemagglutination is an unreliable routine method for Lewis antigen designation. Now genomic typing of the Lewis gene is available. Additionally, flow cytometry has been used for typing. We wanted to compare the results of Lewis typing in healthy individuals using the three methods. Materials and Methods: Ninety–three randomly selected plasma donors were genotyped for inactivating Secretor (FUT2) G428A and Lewis (FUT3) T59G, T202C, C314T, G508A and T1067A point mutations. All Le(a+b–) individuals (nonsecretors) were homozygous for the FUT2 G428A mutation and all Le(a–b–) individuals had inactivating mutations on both FUT3 alleles. Fixed erythrocytes were analyzed by fluorescence–activated flow cytometry and the results were compared with hem– agglutination and genotypic data. Antigen availability was expressed as median fluorescence intensity and as percentage positive cells with fluorescence intensities ≥102. Results: Using an anti–Lea reagent a mean of 99% of erythrocytes from Le(a+b–) individuals and 1% of erythrocytes from Le(a–b–) or Le(a–b+) individuals were stained positive. Using an anti–Leb reagent, a mean of 71% of erythrocytes from A1, 95% from B and 99% from O and A2 Le(a–b+) individuals and less than 10% of erythrocytes from Le(a–b–) or Le(a+b–) individuals were stained positive. After papain treatment 100% of the erythrocytes from A1 and A1B Le(a–b+) individuals stained positive without increase in background staining. The flow cytometric technique revealed large differences in staining intensities, within each ABO Le(a–b+) subgroup which was not directly correlated to plasma donation frequencies nor to Secretor or Lewis genotypes. Conclusion: Flow cytometry may prove valuable as a Lewis blood group typing technique but also as a research tool when investigating Lewis phenotypes of human erythrocytes.

1.
Henry S, Oriol R, Samuelsson B: Lewis histo–blood group system and associated secretory phenotypes. Vox Sang 1995;69:166–182.
2.
Schenkel–Brunner H: Human Blood Groups: Chemical and Biochemical Basis of Antigen Specificity. Wien, Springer, 1995.
3.
Oriol R: ABO, Hh, Lewis and Secretion – Serology, Genetics and Tissue Distribution; In Blood Cell Biochemistry. New York, Plenum Press, 1995, vol 6: Molecular Basis of Major Human Blood Group Antigens (edited by Cartron JP, Rouger P), pp 37–73.
4.
Mollicone R, Cailleau A, Oriol R: Molecular genetics of H, Se, Lewis and other fucosyltransferase genes. Transfus Clin Biol 1995; 4:235–242.
5.
Kukowska–Latallo J, Larsen R, et al.: A cloned human cDNA determines expression of a mouse stage–specific embryonic antigen and the Lewis blood group α(1,3/1,4)fucosyltransferase. Genes Dev 1990;4:1288–1303.
6.
Kelly RJ, Rouquier S, Giorgi D, et al: Sequence and expression of a candidate for the human Secretor blood group α(1,2)fucosyltransferase gene (FUT2) – Homozygosity for an enzyme–inactivating nonsense mutation commonly correlates with the nonsecretor phenotype. J Biol Chem 1995;270:4640–4649.
7.
Elmgren A, Rydberg L, Larson G: Genotypic heterogeneity among Lewis negative individuals. Biochem Biophys Res Commun 1993;196: 515–520.
8.
Nishihara S, Yazawa S, Iwasaki H, et al.: α(1,3/1,4)Fucosyltransferase (FucT–III) gene is inactivated by a single amino acid substitution in Lewis histo–blood type negative individuals. Biochem Biophys Res Commun 1993; 196:624–631.
9.
Koda Y, Kimura H, Mekada E: Analysis of Lewis fucosyltransferase genes from the human gastric mucosa of Lewis–positive and Lewis–negative individuals. Blood 1993;82: 2915–2919.
10.
Mollicone R, Reguigne I, Kelly RJ, et al: Molecular basis for Lewis α(1,3/1,4)fucosyltransferase gene deficiency (FUT3) found in Lewis–negative Indonesian pedigrees. J Biol Chem 1994;269:20987–20994.
11.
Nishihara S, Narimatsu H, Iwasaki H, et al: Molecular genetic analysis of the human Lewis histo–blood group system. J Biol Chem 1994;269:29271–29278.
12.
Elmgren A, Börjeson C, Svensson L, et al: DNA sequencing and screening for point mutations in the human Lewis (FUT3) gene enables molecular gentoyping of the human Lewis blood group system. Vox Sang 1996;70: 97–103.
13.
Yu LC, Yang YH, Broadberry RE, et al: Correlation of a missense mutation in the human Secretor α1,2fucosyltransferase gene with the Lewis(a+b+) phenotype: A potential molecular basis for the weak Secretor allele (Se–W). Biochem J 1995;312:329–332.
14.
Henry S, Mollicone R, Lowe JB, et al: A second non secretor allele of the blood group α(1,2)fucosyltransferase gene (FUT2). Vox Sang 1996;70:21–25.
15.
Yu LC, Broadberry RE, Yang YH, et al: Heterogeneity of the human Secretor α(1, 2)fucosyltransferase gene among Lewis (a+b–) non secretors. Biochem Biophys Res Commun 1996;222:390–394.
16.
Henry S, Mollicone R, Fernandez P, et al: Molecular basis for erythrocyte Le(a+b+) and salivary ABH partial–secretor phenotypes. Expression of a FUT2 secretor allele with an A→T mutation at nucelotide 385 correlates with reduced α(1,2)fucosyltransferase activity. Glycoconj J 1996;13:985–993.
17.
Ørntoft TF, Vestergaard EM, Holmes E, et al: Influence of Lewis α1–3/4–L–fucosyltransferase (FUT3) gene mutations on enzyme activity, erythrocyte phenotyping, and circulating tumor marker sialyl–Lewis a levels. J Biol Chem 1996;271:32260–32268.
18.
Elmgren A, Mollicone R, Costache M, et al: Significance of individual point mutations, T202C and C314T, in the human Lewis (FUT3) gene for expression of Lewis antigens by human α(1,3/1,4)fucosyltransferase, FUC–TIII. J Biol Chem 1997;272:21994–21998.
19.
Narimatsu H, Iwasaki H, Nakayama F, et al: Lewis and secretor gene dosage affect CA19–9 and Du–PAN–2 serum levels in normal individuals and colorectal patients. Cancer Res 1998; 58:512–518.
20.
Sneath J, Sneath P: Transformation of the Lewis groups of human red cells. Nature 1955; 176:172.
21.
Marcus D, Cass L: Glycosphingolipids with Lewis blood group activity: Uptake by human erythrocytes. Science 1969;164:533–555.
22.
Hammar L, Månsson S, Rohr T, et al: Lewis phenotype of erythrocytes and Leb–active glycolipid in serum of pregnant women. Vox Sang 1981;40:27–33.
23.
Langkilde NC, Wolf H, Ørntoft TF: Lewis antigen expression in benign and malignant tissues from RBC Le(a–b–) cancer patients. Br J Haematol 1991;79:493–499.
24.
Stigendal L, Olsson R, Rydberg L, Samuelsson BE: Blood group Lewis phenotype on erythrocytes and in saliva in alcoholic pancreatitis and chronic liver disease. J Clin Pathol 1984;37: 778–782.
25.
Karlsson K–A, Larson G: Structural characterization of lactotetraosylceramide, a novel glycosphingolipid isolated from human meconium. J Biol Chem 1979;254:9311–9316.
26.
Kudo T, Iwasaki H, Nishihara S, et al: Molecular genetic analysis of the human Lewis histo–blood group system. 2. Secretor gene inactivation by a novel single missense mutation A385T in Japanese non secretor individuals. J Biol Chem 1996;271:9830–9837.
27.
Hanfland P: Isolation and purification of Lewis blood group active glycosphingolipids from the plasma of human O Leb individuals. Eur J Biochem 1978;87:161–170.
28.
Borén T, Falk P, Roth KA, et al: Attachment of Helicobacter pylori to human gastric epithelium mediated by blood group antigens. Science 1994;262:1892–1895.
29.
Ilver D, Arnqvist A, Ögren J, et al: Helicobacter pylori adhesin binding fucosylated histo–blood group antigens revealed by retagging. Science 1998;279:373–377.
30.
Murai J, Naka K, Shimojo N, et al: Quantitative analysis of Lewis antigens on erythrocytes by flow cytometry. Clin Chim Acta 1994; 226:21–28.
31.
Mollison P, Engelfriet C, Contreras M: In Blood Transfusion in Clinical Medicine, ed 10. Oxford, Blackwell Scientific Publications, 1997.
32.
Good AH, Yau O, Lamontagne L, Oriol R: Serological and chemical specificities of twelve monoclonal anti–Lea and anti–Leb antibodies. Vox Sang 1992;62:180–189.
33.
Pisacka M, Stambergova M: Activation of Thomsen–Friedenreich antigens on red cells: A possible source of errors in antigen typing with some monoclonal antibodies. Vox Sang 1994;66:300.
34.
Box GEP, Hunter WG, Hunter J: In Statistics for Experimenters. New York, Wiley, 1978.
35.
Haaland PD: In Experimental Design in Biotechnology. New York, Dekker, 1989.
36.
Karlsson KA: Preparation of total nonacid glycolipids for overlay analysis of receptors for bacteria and viruses and for other studies. Methods Enzymol 1987;138:212–220.
37.
Magnani JL, Smith DF, Ginsburg V: Detection of gangliosides that bind cholera toxin: Direct binding of 125I–labeled toxin to thin–layer chromatograms. Anal Biochem 1981;109:399–402.
38.
Brockhaus M, Magnani J, Blaszczyk M, et al: Monoclonal antibodies against the human Leb blood group antigen. J Biol Chem 1981;256: 13223–13225.
39.
Hansson GC, Karlsson KA, Larson G, et al: Detection of blood group type glycosphingolipid antigens on thin–layer plates using polyclonal antisera. J Immunol Methods 1985; 83:37–42.
2000
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