Oligosaccharide differences in the DF3 sialomucin antigen from normal human milk and the BT-20 human breast carcinoma cell line

The murine monoclonal antibody DF3, prepared against a membrane-enriched fraction of a human breast carcinoma, recognized high molecular weight mucin-like glycoproteins from normal human milk and breast carcinoma cell lines. Although the epitope recognized appeared to be a peptide segment, recognition was altered by neuraminidase, suggesting carbohydrate contributions to antigen recognition. Examination of DF3 antigen isolated from normal human milk and the BT-20 human breast carcinoma cell line showed significant oligosaccharide differences. DF3 antigen from BT-20 cells contained three major oligosaccharides, the peanut agglutinin-binding disaccharide Gal beta 1,3GalNAc, which is the carbohydrate component of the Thomsen-Friedenreich antigen, and its mono-(NeuAc2,3Gal beta 1,3GalNAc and/or Gal beta 1,3(NeuAc alpha 2,6)GalNAc) and disialylated (NeuAc2,3Gal beta 1,3(NeuAc alpha 2,6)GalNAc) derivatives. In contrast, the DF3 antigen from normal human milk contained the tetrasaccharide Gal beta 1,4GlcNASc beta 1,6(Gal beta 1,3)GalNAc as its major neutral oligosaccharide and also sialylated derivatives of the tetrasaccharide, including a monosialo derivative (Gal beta 1,4GlcNac beta 1,6(NeuAc alpha 2,3Gal beta 2,3)GalNAc and/or NeuAc alpha 2,3Gal beta 1,4GlcNAc beta 1,6 (Gal beta 1,3)GalNAc). These results suggest the possibility of carcinoma-associated alterations in O-linked oligosacchardes of cell surface sialomucins and in the activity of the beta 1,6-glucosaminyl transferase involved in mucin biosynthesis.     

Comparative study of the N-linked oligosaccharides released from normal human esophageal epithelium and esophageal squamous carcinoma

N-Linked sugar chains of normal human esophageal epithelium and esophageal squamous carcinoma were quantitatively released as oligosaccharides from their membrane preparations by hydrazinolysis. After being fractionated by serial lectin column chromatography using concanavalin A-Sepharose and Datura stramonium agglutinin-Sepharose, their structures were elucidated by exoglycosidase digestion in combination with methylation analysis. Both normal epithelium and esophageal carcinoma contained bi-, tri- and tetraantennary oligosaccharides as well as high mannose-type oligosaccharides. Interestingly, carcinoma had about 1.6 times larger amounts of tri- and tetraantennary oligosaccharides with the GlcNAc beta 1----4Man alpha 1----and/or the GlcNAc beta 1----6Man alpha 1----linkages than normal epithelium. Tri- and tetraantennary oligosaccharides with N-acetyllactosamine repeating units (the Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4GlcNAc group) were also increased in carcinoma. These data indicated that the altered glycosylation of proteins previously found in transformed rodent cells also occurs widely in human esophageal carcinoma.     

Structures of the asparagine-linked oligosaccharides of an alkaline phosphatase, kasahara isozyme, purified from FL amnion cells

Asparagine-linked oligosaccharides were quantitatively released by hydrazinolysis from an alkaline phosphatase, Kasahara isozyme, which was purified from FL amnion cells. Almost all of the oligosaccharides (98%) were acidic components, all of which can be converted to neutral oligosaccharides upon sialidase digestion. Structural analysis of the oligosaccharides by sequential exoglycosidase digestion in combination with methylation analysis revealed that the alkaline phosphatase of FL cells contains sialylated mono-, bi-, tri-, and tetraantennary complex type sugar chains with the Gal beta 1----4GlcNAc beta 1---- outer chains. Some of the tetraantennary sugar chains contain a single Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4GlcNAc beta 1---- outer chain on their Man alpha 1----6 arm. Both fucosylated and nonfucosylated trimannosyl cores were found in the sugar chains. However, it is of interest that the core portion of monoantennary oligosaccharide was not fucosylated and that of the tetraantennary oligosaccharide with a tetrasaccharide outer chain was completely fucosylated.     

Glycoproteins associated with metastatic potential of cancer

Carbohydrate moieties of glycoproteins have been implicated to be involved in cellular adhesion. Therefore, certain carbohydrate structures in glycoproteins are expected to be associated with metastatic behaviour of cancer cells; such carbohydrate structure can be used as an indicator for the treatment of cancer patients based on the predicted metastatic potential of the cancer cells. The following results recently reported is interesting from the above view point. In certain cancer cells of the mouse, beta 1----6 branching of asparagine-linked oligosaccharides, which can be detected by the reactivity with PHA-L lectin, associates with increased metastatic potential. In human bladder carcinomas, reactivity with Lotus tetragonolobus agglutinin (LTA) correlates with increased metastatic potential. LTA appears to react with Lex structure [Gal beta 1----4(Fuc alpha 1----3)GlcNAc] in the cancer cells. Expression of sialilated dimeric Lex increases in the metastatic nests of human colon carcinomas. On the other hand, sialyl antigen MGl has the tendency to be expressed in human gastric adenocarcinoma of low metastatic potential. MGl is defined by a monoclonal antibody raised against Ricinus communis agglutinin receptors isolated from human gastric adenocarcinoma xenografted in nude mice. Continued efforts using monoclonal antibodies and lectins may yield arrays of carbohydrate markers of clinical value to predict the metastatic potential.     

Carbohydrate analysis of immunoglobulin G myeloma proteins by lectin and high performance liquid chromatography: role of glycosyltransferases in the structures

The carbohydrate structures and the enzymatic basis for glycosylation of IgG by bone marrow plasma cells were determined in 7 patients with monoclonal gammopathy of undetermined significance and 22 patients with IgG MM. Lectin-binding analysis showed that in all cases of monoclonal gammopathy of undetermined significance and normal controls the IgG heavy chains bound to Ricinus communis agglutinin more strongly than to concanavalin A. In contrast, the IgG in 11 of the 17 advanced cases of MM (stages II and III) studied reacted to concanavalin A more strongly. Structural analysis showed that the reduced R. communis agglutinin binding capacity of these MM IgGs was due to hypogalactosylation of IgG. The galactosyltransferase and N-acetylglucosaminyltransferase III activities of the bone marrow myeloma cells from 5 MM cases were found to have a low enzyme activity ratio of galactosyltransferase to N-acetylglucosaminyltransferase III which reflects the hypogalactosylation. This indicates that the difference in the carbohydrate moieties observed in myeloma proteins is due to variations in the activities of the two glycosyltransferases.     

Structural studies of the carbohydrate moieties of carcinoembryonic antigens

Three samples of carcinoembryonic antigens were purified from liver metastases of primary colon cancer. The asparagine-linked sugar chains of carcinoembryonic antigens (CEA) were released as oligosaccharides by hydrazinolysis and the structures of oligosaccharides, thus obtained, was studied in combination with methylation analysis and several limited exoglycosidase digestions. All three CEAs contain approximately 25 asparagine-linked sugar chains in one molecule and about 10% of them was high mannose type. However, structural features of the outer chain moieties of the remaining complex-type sugar chains were different by CEA samples. The complex-type sugar chains were mono-, bi-, tri-, and tetraantennary with Man alpha 1----6(+/- GlcNAc beta 1----4)(Man alpha 1----3)Man beta 1----4GlcNAc beta 1----4(+/- Fuc alpha 1----6)GlcNAc as their cores, half of which were bisected; 86% of their proximal N-acetylglucosamine was fucosylated. The major outer chains in two samples were N-acetyllactosamine and Gal beta 1----4(Fuc alpha 1----3)GlcNAc (X-antigenic determinant) and the remaining one sample contained Fuc alpha 1----2Gal beta 1----4(Fuc alpha 1----3)GlcNAc (Y-antigenic determinant) as an additional major outer chain. Furthermore, small amounts of type 1 chain and Lea antigenic determinant were found in some samples. Acidic oligosaccharides consisted of sialic acid containing fractions and sialidase-resistant fractions, and their contents seemed to be in a reciprocal relationship. Sialic acid was linked at the C-3 and C-6 positions of the nonreducing terminal galactose residues of the outer chains.     

Structure of the sugar chain of alphafetoprotein purified from a human yolk sac tumor and its reactivity with concanavalin A

We have attempted to determine the carbohydrate moiety of human alphafetoprotein (AFP) produced by a yolk sac tumor. AFP was obtained from the cystic fluid of human yolk sac tumors grown in nude mice and was purified using an immunoadsorbent column coupled with monoclonal anti-AFP antibody. Then, the carbohydrate chain of the purified AFP was quantitatively released from the polypeptide chain. The resulting oligosaccharide was labeled and, by sequential exoglycosidase digestion in combination with methylation analysis and periodate oxidation, the structure was determined to be: Sia alpha 2----6Gal beta 1----4GlcNAc beta 1----2Man alpha 1----6 (GlcNAc beta 1----4) (Sia alpha 2----6Gal beta 1----4GlcNAc beta 1----2Man alpha 1----3) Man beta 1----4GlcNAc beta 1----4 (Fuc alpha 1----6) GlcNAcOT Compared with the known structure of the sugar, chain of human hepatic AFP, it was found that the sugar chain of yolk sac AFP contained an additional sugar, N-acetylglucosamine (bisect GlcNAc) linked to the beta-mannose. In the light of recent knowledge, this result indicates that the Concanavalin A (Con-A) binding site of the sugar chain is blocked by this GlcNAc in human yolk sac AFP. This fact forms the basis for the clinical use of the Con-A binding test to determine the origin of AFP in patients.     

Structure of the asparagine-linked sugar chains of alpha-fetoprotein purified from human ascites fluid

alpha-Fetoprotein purified from human serum was found to contain an asparagine-linked sugar chain in one molecule. The sugar chain was quantitatively released from the polypeptide moiety by hydrazinolysis and recovered as oligosaccharides after N-acetylation. The oligosaccharide mixture was separated into a neutral (N) and two acidic (A-1 and A-2) fractions by paper electrophoresis. By combination of sequential exoglycosidase digestion, methylation analysis, and concanavalin A-Sepharose column chromatography, the structures of these fractions were determined to be: Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GLcNAc beta 1 leads to 4(+/- Fuc alpha 1 leads to 6)GlcNAc; Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(+/- Fuc alpha 1 leads to 6(GlcNAc; and NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 6(NeuAc alpha 2 leads to 6Gal beta 1 leads to 4GlcNAc beta 1 leads to 2Man alpha 1 leads to 3)Man beta 1 leads to 4GlcNAc beta 1 leads to 4(+/- Fuc alpha 1 leads to 6)GlcNAc.     

Modification of cell surface carbohydrates and invasive behavior by an alkyl lysophospholipid

The effect of the alkyl lysophospholipid racemic-1-O-octadecyl-2-O-methyl glycero-3-phosphocholine on the expression of cell surface carbohydrates of four matched pairs of normal and malignant cells was studied using chromatographic techniques. After treatment with alkyl lysophospholipid, glycopeptides proteolytically derived from normal and malignant cells displayed a shift in the size distribution profiles obtained by gel filtration. These drug-induced changes in molecular weight distribution were expressed most strongly in untransformed cells and resembled the carbohydrate alterations found after their malignant transformation. Desialylation abolished the effect of alkyl lysophospholipid, thus suggesting an increased amount of sialic acid in the surface carbohydrates of drug-treated cells. Chromatography of glycopeptides on concanavalin A-Sepharose, Ricinus communis agglutinin I-agarose, and Bio-Gel P-4 columns excluded a higher degree of branching but suggested addition of extra terminal sialic acid residues as the major cause of the observed alterations. Alkyl lysophospholipid stimulated glycoprotein sialylation of normal cells to the level observed in malignant cells, thus inducing a "malignant-like" surface phenotype. The drug-induced carbohydrate changes in normal chick heart tissue prevented its being invaded by tumor cells when tested in an organotypic assay. The alkyl lysophospholipid thus appears to modulate in a nontoxic fashion the expression of surface molecules implicated in various cellular interactions including invasiveness.     

Characterization of mucin antigens recognized by monoclonal antibodies raised against human colon cancer cells

Two monoclonal antibodies, MLS 102, which recognizes cancer-associated mucin antigens, and MLS 103, which recognizes normal mucin, were used to isolate, by immunoaffinity chromatography, the corresponding antigens from cell lysates and spent medium of a human colorectal carcinoma cell line, LS 180. The MLS 102 antigen contained serine, threonine, and proline as major amino acids. The carbohydrate chains of the MLS 102 antigen were composed of O-linked NeuAc alpha 2----6GalNAc (56%), N-acetylgalactosamine (25%), and longer oligosaccharide chains. The MLS 103 antigen differed from the MLS 102 antigen in both amino acid and carbohydrate composition. Most O-linked oligosaccharides of the MLS 103 antigen were longer than the disaccharide found in the MLS 102 antigen. Immunostaining of LS 180 cells using MLS 102 and MLS 103 revealed that the cells are heterogeneous with respect to the expression of the antigens.     

Tissue binding of lectins in disorders of the breast

Twenty breast lesions including seven scirrhous ductal carcinomas, one infiltrating lobular carcinoma, one colloid carcinoma, four fibroadenomas, and seven cases of fibrocystic disease were analyzed by fluorescence microscopy for the presence and distribution of lectin-binding carbohydrates. Paraffin-embedded tissue sections were tested with wheat germ agglutinin (WGA), Ricin communis agglutinin I (RCA I), peanut agglutinin (PNA), Soybean agglutinin (SBA), Dolichos biflorus agglutinin (DBA), Ulex europaeus agglutinin I (UEA I), and concanavalin A (Con A). Brightest and most consistent staining regardless of the nature of the breast lesion was obtained with WGA followed in approximate order of staining intensity by RCA, PNA, SBA/DBA and Con A. UEA I stained many of the benign breast lesions but no malignant lesions. Lectin binding carbohydrate in benign lesions was localized mainly along the apices of mammary epithelial cells but there was considerable variation in staining patterns among malignant tumors. The fluorescence microscopic arrangement of lectin binding carbohydrate appears distinct for each malignant neoplasm of breast but is more consistent in benign conditions.     

Comparative Study of the N-Linked Oligosaccharides Released from Normal Human Esophageal Epithelium and Esophageal Squamous Carcinoma

N -Linked sugar chains of normal human esophageal epithelium and esophageal squamous carcinoma were quantitatively released as oligosaccharides from their membrane preparations by hydrazinolysis. After being fractionated by serial lectin column chromatography using concanavalin A-Sepharose and Datura stramonium agglutinin-Sepharose, their structures were elucidated by exoglycosidase digestion in combination with methylation analysis. Both normal epithelium and esophageal carcinoma contained bi-, tri- and tetraantennary oligosaccharides as well as high mannose-type oligosaccharides. Interestingly, carcinoma had about 1.6 times larger amounts of tri- and tetraantennary oligosaccharides with the GlcNAcβ1→4Manα1→ and/or the GlcNAcβ1→6Manα1→ linkages than normal epithelium. Tri- and tetraantennary oligosaccharides with N -acetyllactosamine repeating units (the Galβ1→4GlcNAcβ1→3Galβ1→4GlcNAc group) were also increased in carcinoma. These data indicated that the altered glycosylation of proteins previously found in transformed rodent cells also occurs widely in human esophageal carcinoma.     

Altered protein glycosylation of rat 3Y1 cells induced by activated c-myc gene

N-linked sugar chains of rat 3Y1 cells and tumorigenic cells derived by transfection with activated c-myc gene were quantitatively released as oligosaccharides from membrane preparations by hydrazinolysis. Structural analyses revealed that cells of both types contain bi-, tri- and tetra-antennary complex-type oligosaccharides as well as high-mannose-type oligosaccharides. However, the c-myc-transfected cells showed an increase in tri- and tetra-antennary oligosaccharides having the GlcNAc beta 1----4Man alpha 1----and/or the GlcNAc beta 1----6Man alpha 1----linkages with a decrease in biantennary oligosaccharides compared to control 3Y1 cells. The data suggest that c-myc gene has a potential role in the regulation of cellular protein glycosylation and that an elevated expression of c-myc gene in the cells leads to increased branch formation of outer chains in N-linked oligosaccharides concomitant with the acquisition of tumorigenicity.     

Oncodevelopmental expression of--GlcNAc beta 1-6Man alpha 1-6Man beta 1--branched asparagine-linked oligosaccharides in murine tissues and human breast carcinomas

Increased--GlcNAc beta 1-6Man alpha 1-6Man beta--branching in asparagine-linked oligosaccharides has been observed in murine and human tumor cells and has recently been linked to enhanced metastatic potential in experimental tumor models. Leukoagglutinin (L-PHA) requires the beta 1-6-linked lactosamine antenna (beta 1-6 branch) for high affinity binding and was used in this study to quantitate these structures on glycoproteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Normal rodent tissues and cell lines were used to standardize the experimental conditions required to quantitate beta 1-6-branched oligosaccharide structures and the glycosyltransferase activity which initiates the synthesis of the antenna, beta 1-6 N-acetylglucosamine (GlcNAc)-transferase V (EC 2.4.1.155). Secondly, the levels of L-PHA-reactive oligosaccharide were compared in a series of benign and malignant human breast biopsies. Normal human breast tissue and benign lesions showed low expression but 50% of the primary malignancies examined showed significantly elevated L-PHA reactivity. GlcNAc transferase V activities in the human breast carcinomas and in normal murine tissues correlated with the levels of L-PHA reactive oligosaccharide in the tissues. GlcNAc transferase V showed similar ranges of activities, differing by approximately 5-fold between high and low expressing mouse tissues; fibroblasts with and without an activated H-ras oncogene; and low and high expressing human breast carcinomas. The results show that beta 1-6 branching in asparagine-linked oligosaccharides is dependent on tissue-specific regulation of GlcNAc transferase V activity. Secondly, a subset of human breast malignancies showed elevated levels of beta 1-6-branched oligosaccharides compared to benign samples, suggesting that further studies are warranted to determine whether the presence of these oligosaccharides is associated with metastatic disease and reduced patient survival time.     

Asn-linked oligosaccharide processing and malignant potential

There is now considerable evidence that malignant transformation is accompanied by many changes in the oligosaccharide structures of glycoproteins and glycolipids. More recently, studies using tumour cell glycosylation mutants and inhibitors of Asn-linked oligosaccharide processing have suggested that the transformation-associated increase in GlcNAc beta 1-6Man alpha 1-6 Man beta- branching of complex-type oligosaccharides may enhance tumour cell metastasis. The synthesis of beta 1-6 branched oligosaccharides and related structures as well as their possible function in cell-cell interactions and tumour cell metastasis are discussed.     

Effect of neuraminidase and papain treatment on lectin-induced agglutination of Novikoff tumor cells and assay of lectin receptor activity of the glycopeptides released from the cell surface by papain.

Lectins, plant proteins that bind specific saccharide determinants, have been utilized to examine the effect of neuraminidase digestion on the structure and/or expression of oligosaccharide moieties present at the periphery of Novikoff ascites hepatoma cells. Five lectins were utilized: concanavalin A (Con A), specific for alpha-D-manno- or alpha-D-glucopyranosyl residues; wheat germ agglutinin, specific for 2-acetamido-2-deoxy-D-glucopyranosyl residues; Ricinus communis agglutinin I (RCAI), specific for D-glucopyranosyl residues; R. communis agglutinin II (RCAII), specific for D-galacto- or 2-acetamido-2-deoxy-D-galactopyranosyl residues; and soybean agglutinin, specific for 2-acetamido-2-deoxy-D-galactopyranosyl residues. Neuraminidase treatment of Novikoff cells did not alter their agglutination by Con A or wheat germ agglutinin. Similar treatment produced only a 2-fold increase in their agglutination by RCAI but a 12-fold increase in their agglutination by RCAII, indicating that 2-acetamido-2-deoxy-D-galactopyranosyl residues become expressed upon neuraminidase treatment. This conclusion was confirmed by the observation that neuraminidase-treated Novikoff cells acquired agglutinability by soybean agglutinin. Binding studies using ferritin-conjugated RCAII indicated that neuraminidase treatment exposed cryptic cell surface receptors for RCAII. To ascertain the role of cell surface glycoproteins in lectin-induced agglutination of Novikoff cells, glycopeptides cleaved from the cell surface by papain were assayed for lectin receptor activity. The cell surface glycopeptides exhibited receptor activity for Con A, wheat germ agglutinin and RCAI but not for RCAII and soybean agglutinin. A cell surface macrosialoglycopeptide fraction, resolved by gel filtration and ion-exchange chromatography, possessed a major portion of the Con A and RCAI receptor activity.     

末端Galβ1-4 GlcNAc 多糖分子在肿瘤患者血清中的表达

 目的 研究末端Galβ1—4GlcNAc多糖分子在鼻咽癌、肺癌、肝癌和结直肠癌患者血清中的表达规律。方法 应用RCAⅠ（蓖麻凝集素Ⅰ）亲和层析法从正常人血清中获得RCAI相关末端Galβ1—4GlcNAC糖蛋白，用辣根过氧化物酶标记，采用凝集素差减法，对133例肿瘤患者，64例良性疾病患者和240例正常人血清进行检测。结果 鼻咽癌、肝癌和结直肠癌患者血清中RCAⅠ识别的多糖分子明显高于正常人和良性疾病患者，阳性率分别为66．7％、73．9％、64．9％；肺癌患者血清阳性率为19．4％。且在鼻咽癌和结直肠癌患者有效治疗后该多糖分子水平显著下降，阳性率也下降。结论 血清中RCAⅠ相关的末端Galβ1—4GlcNAc多糖分子，可用于肿瘤患者血清糖基化异常和肿瘤发病机制的研究，并可为临床诊断和治疗效果的评价提供参考指标。     

A carbohydrate epitope associated with human squamous lung cancer

A monoclonal antibody, 43-9F, specifically recognizes a tumor-associated antigen expressed both on surface membrane glycoproteins and on secreted soluble mucins of human squamous lung carcinoma (SLC) cells, and the corresponding antigen can be detected as a circulating tumor marker in plasma of SLC patients. Thin-layer chromatography immunostaining of neutral glycolipids extracted from SLC cells reveals a 43-9F-reactive glycolipid whose carbohydrate structure, as determined by fast atom bombardment-mass spectrometry, is identical with that of an Lea-active pentaglycosylceramide described previously: Gal beta 1-3[Fuc alpha 1-4]-GlcNAc beta 1-3Gal beta 1-4Glc-Cer. However, the Lea-active oligosaccharide hapten, lacto-N-fucopentaose II, with the same carbohydrate structure, fails to inhibit binding of 43-9F, and a well-characterized anti-Lea monoclonal antibody blocks only 40% of 43-9F binding sites on SLC cells, suggesting that the major epitope recognized by 43-9F is more complex than the Lea epitope. To search for a higher affinity 43-9F epitope among more complex oligosaccharides, a mixture of tritiated neutral oligosaccharide alditols from pooled human milk was passed through a 43-9F affinity column. A major retarded oligosaccharide was purified by high-performance liquid chromatography and shown by fast atom bombardment-mass spectrometry to have the following structure: Gal beta 1-3[Fuc alpha 1-4]GlcNAc beta 1-3Gal beta 1-4[Fuc alpha 1-3] GlcNAc beta 1-3Gal beta 1-4Glc. Oligosaccharides containing this sugar sequence are at least 100-fold more active than lacto-N-fucopentaose II as competitive inhibitors of 43-9F. Thus, antibody 43-9F binds to the above difucosyl Lea-X determinant with high affinity and weakly cross-reacts with the Lea antigen under some conditions such as occurs in thin-layer chromatography and enzyme-linked immunosorbent assay where multiple weak interactions of the decavalent IgM antibody may occur.     

Cell surface asparagine-linked sugar chains of human early myeloblastic leukemic cells (KG-1a)

The asparagine-linked sugar chains obtained from total cell surface membrane glycoproteins of human early myeloblastic leukemic cells (KG-1a cells) were studied. The sugar chains liberated by hydrazinolysis were purified by paper electrophoresis, paper chromatography, and Bio-Gel P-4 chromatography followed by analysis of exoglycosidase digestion and methylation study. Neutral oligosaccharides were all composed of high mannose type sugar chains. Acidic oligosaccharides were chiefly composed of typical bi-, tri-, and tetraantennary complex type sugar chains with Gal beta 1----4GlcNAc beta 1----groups and Neu-Ac alpha 2----3 or 6Gal beta 1----4GlcNAc beta 1----groups (in which Gal is galactosyl, GlcNac is N-acetylglucosamine, and NeuAc is N-acetylneuraminic acid) as side chains. Moreover the following two structures were identified in (in which Fuc is fucosyl): monosialyl bi- and triantennary sugar chains with a Gal beta 1----4(Fuc alpha 1----3)GlcNAc beta 1----group (X determinant) as one of the side chains; and monosialyl tetraantennary sugar chains with a Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4GlcNAc group (repeating N-acetyllactosamine unit) as one of the side chains. These data together with our previous studies on sugar chains of K562 cells [early erythroblast], adult erythrocytes [H. Yoshima, N. Shiraishi, A. Matsumoto, S. Maeda, T. Sugiyama, and A. Kobata, J. Biochem. (Tokyo), 91: 233-246, 1982], and HL-60 cells [promyelocyte] [A. Mizoguchi, S. Takasaki, S. Maeda, and A. Kobata, J. Biol. Chem., 259: 11943-11957, 1984] strongly suggest that the cell surface asparagine-linked sugar chains alter in an orderly fashion, systematically in association with lineage and maturation stages during hematopoietic cell differentiation.