M1 Antigens Research Articles

Mechanisms of HLA-DR antigen expression in phytohemagglutinin-activated T cells in man. Requirement of T cell recognition of self HLA-DR antigen expressed on the surface of monocytes.

Signals required for expression of HLA-DR (DR) antigen in phytohemagglutinin (PHA)-activated human peripheral blood T cells were examined. T cells were purified by a four-step procedure, which included depletion of glass-adherent cells, 53% Percoll gradient centrifugation, nylon wool column passage, and treatment with mouse monoclonal antibodies directed to human HLA-DR antigen and Leu M1 antigen plus complement. Purified T cells responded poorly to PHA but with the combination stimuli of PHA and recombinant human interleukin 2 (rIL-2), resting T cells proliferated as well as T cells cultured with 10% monocytes and PHA. But well proliferated T cells in the absence of monocytes expressed very poor DR antigen after 7 to 8 days of culture. DR expression of T cells was restored by the addition of 10% monocytes. Allogeneic monocytes also helped proliferative responses of PHA-activated T cells but did not help the expression of DR antigen. These results suggested that signals required for T cell proliferation (PHA and rIL-2) were not sufficient for DR expression in this system and further monocytes were essentially required in a HLA-restricted manner. In the next experiment, we examined the role of membrane molecules in monocytes for transmission of signals that induce activated T cells to express DR antigen. Autologous monocytes were fixed with 1% paraformaldehyde and added to T cells in the presence of PHA and rIL-2. Fixed monocytes could help DR antigen expression of PHA-activated T cells as well as viable monocytes. But when fixed monocytes were pretreated with anti-HLA-DR monoclonal antibody, they could not help DR expression of T cells any longer. These results suggested that for the expression of DR antigen, PHA-activated T cells had to first recognize self DR antigen expressed on the surface of monocytes before proliferation occurred.

Establishment of a common acute lymphoblastic leukemia cell line (LC4-1) and effects of phorbol myristate acetate (PMA) on the surface antigen expression of the cell line.

A common acute lymphoblastic leukemia (ALL) cell line, designated LC4-1, was established from peripheral blood mononuclear cells of a patient with acute non-T-cell ALL. LC4-1 cells were characteristically positive for Ia, B4, and common ALL antigens (CALLA), but negative for B2, Tac, T3, T4, T8, T11, and M1 antigens and E-rosette formation. Approximately 30% of LC4-1 cells expressed detectable amounts of B1 antigens. LC4-1 cells expressed neither Epstein-Barr-virus-associated nuclear antigen (EBNA), cytoplasmic immunoglobulins (cIg) nor surface immunoglobulins (sIg). Gene rearrangements had already occurred in LC4-1 in the D-J region of immunoglobulin heavy chain genes, but not in T-cell receptor (beta-chain) genes, suggesting that LC4-1 is a progenitor cell line of B-cell lineage earlier than pre-B-cells. The expression of cell surface antigens of LC4-1 was changed by treatment with 4-phorbol 12-myristate 13-acetate (PMA) (0.1 ng/ml) for 2 days. Before treatment with PMA, about 98% of LC4-1 cells were positive for B4, CALLA, and Ia. However, following treatment they lost CALLA expression without any change in expression of Ia and B4. There was no change in B1-positive population. The change in surface antigens on LC4-1 cells seems to be due to differentiation induced in the cells by PMA. These results support the hypothesis that CALLA is a differentiation antigen and suggest one possible differentiation pathway for pre-B-cells.

Hexabrachion proteins in embryonic chicken tissues and human tumors.

Cell cultures of chicken embryo and human fibroblasts produce a large extracellular matrix molecule with a six-armed structure that we called a hexabrachion (Erickson, H. P., and J. L. Iglesias, 1984, Nature (Lond.), 311:267-269. In the present work we have determined that the myotendinous (M1) antigen described by M. Chiquet and D. M. Fambrough in chicken tissues (1984, J. Cell Biol., 98:1926-1936), and the glioma mesenchymal extracellular matrix protein described by Bourdon et al. in human tumors (Bourdon, M. A., C. J. Wikstrand, H. Furthmayr, T. J. Matthews, and D. D. Bigner, 1983, Cancer Res. 43:2796-2805) have the structure of hexabrachions. We also demonstrate that the M1 antigen is present in embryonic brain, where it was previously reported absent, and have purified hexabrachions from brain homogenates. The recently described cytotactin (Grumet, M., S. Hoffman, K. L. Crossin, and G. M. Edelman, 1985, Proc. Natl. Acad. Sci. USA, 82:8075-8079) now appears to be identical to the chicken hexabrachion protein. In a search for functional roles, we looked for a possible cell attachment activity. A strong, fibronectin-like attachment activity was present in (NH4)2SO4 precipitates of cell supernatant and sedimented with hexabrachions in glycerol gradients. Hexabrachions purified by antibody adsorption, however, had lost this activity, suggesting that it was due to a separate factor associated with hexabrachions in the gradient fractions. The combined information in the several, previously unrelated studies suggests that hexabrachions may play a role in organizing localized regions of extracellular matrix. The protein is prominently expressed at specific times and locations during embryonic development, is retained in certain adult tissues, and is reexpressed in a variety of tumors.

Human breast cancer cells share antigens with the myeloid monocyte lineage.

We have examined the expression of several myeloid cell associated antigens, some of which are involved in myelomonocyte adhesion, in seven well characterized human breast cancer cell lines, since common properties of adhesiveness and migration are found in haemopoietic cells and epithelial cancer cells. Five of these cell lines were of metastatic origin and two were derived from primary breast carcinoma. Antigenic expression was evaluated by immunofluorescence (IF), flow cytometry (FCM), radioimmunoassay on live cells (RIA) and immunoperoxidase staining. None of these cell lines expressed T or B lymphoid specific antigens. Myeloid antigens My4, MO1, and MOF11 (derived from the hybridization of mouse X63 - Ag8 cells with spleen cells from Balb/c mice immunized with purified human monocytes) were expressed in the 7 cell lines. Leu M1, Leu M3, My9, and MO2 antigens were expressed in some of the cell lines. Leu M2 and My7 antigens were not expressed or at very low levels. The expression of these myeloid antigens was also tested by immunoperoxidase staining, and found on frozen sections of normal mammary gland, fibroadenoma of the breast, primary breast cancer, and lymph node and skin metastases of breast tumours. This common expression in epithelial breast cells and in myeloid cells might be related to common biological functions such as interaction with extracellular matrix which precedes cell migration, a normal function of macrophages and an abnormal function expressed or amplified in human cancer epithelial cells.

Antigenic phenotype of malignant mesotheliomas and pulmonary adenocarcinomas. An immunohistologic analysis demonstrating the value of leu M1 antigen

To evaluate the usefulness of an immunohistologic approach to the differential diagnosis of mesothelioma and pulmonary adenocarcinoma, the authors studied paraffin-embedded, fixed tissue sections from 50 primary adenocarcinomas of the lung and 28 mesotheliomas of the pleura by using a panel of monoclonal antikeratin, antihuman milk fat globule (HMFG-2), anti-Leu M1, and monoclonal anticarcinoembryonic antigen (CEA) antibody; we also used a conventional heterologous anti-CEA antiserum with and without prior absorption with spleen powder to remove antibodies to nonspecific cross-reacting antigen (NCA). Keratin was present in both mesotheliomas and adenocarcinomas and did not help in distinguishing between these two neoplasms. HMFG-2 was detected in 48 (96%), and Leu M1 was positive in 47 (94%) of the adenocarcinomas, but not in any of the mesotheliomas. By using conventional rabbit antiserum, the authors detected CEA in the majority of adenocarcinomas (96%), but also in two cases of mesothelioma. When the anti-CEA antiserum was absorbed with NCA, the number of positively reacting adenocarcinomas decreased considerably to 76%; however, after this treatment, none of the mesotheliomas gave positive reactions. The monoclonal anti-CEA antibody was reactive in 36 of the adenocarcinomas (72%), but in none of the mesotheliomas. Our results indicate that, in addition to HMFG-2 and CEA, the expression of Leu M1 antigen by most primary pulmonary adenocarcinoma (94%) and its absence in mesothelioma could be used as a valuable marker for primary adenocarcinoma of the lung that involves the pleura and permits its differentiation from mesothelioma.

Atypical fibroxanthoma of the skin: a clinicopathological and immunohistochemical study and a discussion of its histogenesis.

The morphological and immunohistochemical characteristics of 37 atypical fibroxanthomas of the skin were examined. Twenty-four tumours were nodular ulcerative lesions on the head and face of patients with a median age of 75 years, whereas 13 tumours occurred on the trunk and limbs of patients with a median age of 48 years. Both pleomorphic polygonal and giant cells as well as the spindle cell component of the tumours stained for the histiocytic markers alpha 1-antichymotrypsin, alpha 1-antitrypsin, lysozyme and, less frequently, for ferritin. Leu M1 antigen and peanut agglutinin receptors were not demonstrable in tumour cells. This antigenic profile was contrasted with the findings in six cases of dermatofibroma which were largely not reactive with the antisera used. The immunohistochemical findings in atypical fibroxanthomas suggest that they represent a homogeneous group of tumours which are related to tissue histiocytes. These results are discussed in the context of the published findings in other so-called fibrohistiocytic tumours including dermatofibrosarcoma protuberans and malignant fibrous histiocytoma. The diagnoses in three cases coded as atypical fibroxanthomas were revised on the basis of their showing a different immunohistochemical profile.

Aneuploidy and expression of gastric-associated mucus antigens M1 and CEA in colorectal adenomas.

A series of 55 flow cytometric characterized colorectal adenomas was analyzed with four different monoclonal antibodies (Mabs) for the occurrence of M1 antigens (associated with gastric fucomucins) and one Mab for carcinoembryonic antigen (CEA). The antigens were detected with an indirect immunoperoxidase technic on paraffin sections after pretreatment with pronase for M1 antigens and without pretreatment for CEA. The staining pattern revealed different correlations with the various parameters, i.e., size, histologic type, atypia, and ploidy of the adenomas. Especially the cytoplasmic staining of anti-M1, Mab (1-13 M1) correlated well with aneuploidy (R = 0.43; P less than or equal to 0.001) and with the DNA index (R = 0.34; P less than or equal to 0.01). Staining of the anti-CEA Mab only correlated with the size of the adenomas (R = 0.29; P less than or equal to 0.03). It is concluded that the immunoreactivity of Mab (1-13 M1), which significantly correlated with aneuploidy, may be associated with malignant transformation in the adenoma-carcinoma sequence.

Primary adenocarcinoma of urinary bladder Clinicopathologic study of 16 cases

The clinicopathologic and immunohistochemical features of 16 pure adenocarcinomas primary in the urinary bladder were reviewed. Only 3 patients were found to have disease confined to the urinary bladder. Of 13 cases with follow-up only 3 are free of disease. Histologically, the tumors were classified as signet ring cell (3), colloid (3), colonic type (5), clear cell (1), and not otherwise specified (NOS, 4). Immunohistochemically, all tumors but one colloid carcinoma were immunoreactive for cytokeratin and epithelial membrane antigen, and most tumors were likewise immunoreactive for carcinoembryonic antigen. Eight cases were immunoreactive for Leu M1 antigen. Prostate specific antigen, S-100 protein, and placental alkaline phosphatase were uniformly negative. No correlation between immunohistochemical profile and histologic type or clinical outcome was found. The utility of immunohistochemistry and other pathologic findings is reviewed.

The M1 and Tm antigens require M and N gene-specified amino acids for expression.

It is known that the major red cell sialoglycoprotein (SGP) of M1 + red cells has the same amino acid sequence as M SGP, whereas that of Tm + cells has the same sequence as N SGP. M1 and Tm are serologically demonstrable when a substitution of N-acetyl-D-glucosamine for N-acetyl-neuraminic acid occurs in one or more of the alkali-labile tetrasaccharides bound at positions 2, 3, and/or 4 of M or N SGP, respectively. The authors used two serums containing anti-M and potent anti-M1 and two containing anti-Tm in tests for antibody crossreactivity and cross adsorption and in antibody inhibition studies. The distinct specificities of anti-M1 and anti-Tm, as determined by these studies, show that the M and N gene-specified amino acids are more important than the glycosylation change in the fine structure of M1 and Tm antigens.

M1 antigens, markers associated with precancerous colonic mucosae: [formula omitted], [formula omitted], [formula omitted]. Laboratoire d'Immunochimie, Unité Cancer et Différenciation Cellulaire ER 277 CNRS. Institut de Recherches Scientifiques sur le Cancer, 94802 Villejuif Cédex, France

M1 antigens, markers associated with precancerous colonic mucosae: [formula omitted], [formula omitted], [formula omitted]. Laboratoire d'Immunochimie, Unité Cancer et Différenciation Cellulaire ER 277 CNRS. Institut de Recherches Scientifiques sur le Cancer, 94802 Villejuif Cédex, France

Use of Leu M1 and antiepithelial membrane antigen monoclonal antibodies for diagnosing Hodgkin's disease.

Biopsies of 82 patients diagnosed as having Hodgkin's disease were reviewed. Seventeen were reclassified histologically as non-Hodgkin's lymphoma or reactive lymphoid hyperplasia. A substantial number of cases of Hodgkin's disease were negative when stained with Leu M1. Staining for Leu M1 was not found in the cases of non-Hodgkin's lymphoma or reactive lymphoid hyperplasia. With the exception of the lymphocyte predominant nodular subtype of Hodgkin's disease, epithelial membrane antigen staining was seen in a few cases of Hodgkin's disease and non-Hodgkin's lymphomas. This was not a useful discriminating feature.

Separation of human B lymphocytes on Helix pomatia A haemagglutinin into two major fractions differing in responsiveness to T-dependent mitogen (pokeweed mitogen) or antigen (tetanus toxoid).

A B-cell fraction consisting of 70% of cells carrying the B-cell-associated B1 antigen, 15-20% of M1+ non-B cells, and less than 3% of T cells was prepared from the peripheral blood of healthy human donors, previously vaccinated with tetanus toxoid (TT). As assessed by immunofluorescence after treatment with neuraminidase, approximately 40-50% of the B cells had surface structures binding to Helix pomatia A haemagglutinin (HP). The cells were separated into three fractions by affinity chromatography on HP conjugated to Sepharose (P, non-retained cells; EI, cells eluted with 0.1 mg/ml N-acetyl-D-galactosamine (D-GalNac); EII, cells eluted with 1 mg D-GalNac/ml). The majority of B cells in fraction EII were HP+ and were rich in cells expressing the B2 differentiation antigen. Sixty per cent of the B cells in this fraction also expressed the major HP-binding glycoprotein, gp 150. In the presence of autologous T cells, these B cells were strongly responsive to activation by either pokeweed mitogen (PWM) or antigen (TT), as reflected by differentiation into plasma cells, secretion of polyclonal IgG and IgM, or IgG anti-TT antibodies. In contrast, fraction P, which contained more than 90% HP-B cells, and which was partially depleted of B2+ cells, responded poorly or not at all to both PWM and TT. Fraction EI was a mixed fraction that responded in an intermediate fashion. When the preparations were depleted of contaminating non-B cells carrying the monocyte or large granular lymphocyte associated M1 antigen, their response to the two stimulating agents did not alter. The results suggest that HP+ B cells differ from HP-B cells in their responsiveness to T-cell signals. Fractionation on unsolubilized HP offers a simple and efficient way of separating B cells into at least two subsets differing in their responsiveness to T-cell-derived differentiation and maturation signals.

Atopic Asthma: T-Cell Response to Corticosteroids

Atopic asthma is associated with diminished cell-mediated immunity and elevated levels of IgE, both of which may be caused by imbalances of T-lymphocyte subsets. We analyzed the response of peripheral blood T-cell subsets to two commonly used corticosteroid preparations as a probe of T-cell subset regulation. We administered prednisone (P) 60 mg or 20 mg, beclomethasone dipropionate (BDP) aerosol, 336 micrograms, placebo, or BDP vehicle in a double-blind protocol to 15 atopic asthmatic patients and ten nonatopic subjects. No difference was found between the groups of the baseline number of T-cells with T4, T8, M1, and Ia antigens, nor the ratio of T4+ (helper) to T8+ (suppressor) cells. Five hours after administration of BDP aerosol, BDP vehicle, and oral placebo, there was no change of these values in either the atopic or in the nonatopic group. In contrast, P, 20 and 60 mg, caused a fall of T4/T8 ratio in the atopic, but not in the nonatopic population. Atopic asthma is not associated with baseline imbalances of peripheral blood T-cell subsets, but is associated with an abnormal response to systemic, but not inhaled corticosteroid.

Maintenance of immunocytologically identified Purkinje cells from mouse cerebellum in monolayer culture

Purkinje cells were identified in monolayer cultures obtained from trypsin-dissociated cerebella of embryonic and early postnatal mice by the Purkinje cell-specific monoclonal antibodies PC1, PC2, PC3 and UCHT1. These cells also expressed the neuronal marker L1 antigen but not the glial markers, glial fibrillary acidic protein or 04 antigen. They also expressed tetanus toxin receptors, PC4, M1 and Thy-1 antigens. Survival of Purkinje cells was best: (a) when cerebella were taken from mice not older than one day of age: (b) when cells were seeded at higher plating densities; and (c) cultured in chemically defined medium which facilitates the survival of neurons. No Purkinje cells could be detected in cultures from mice older than 6 days. PC1 antigen expression developed in vitro on the same time scale as in vivo, i.e. it was first detectable at the equivalent of postnatal days 3–4. At this stage cell bodies had a size of 13–14 μm in diameter and few processes. Dendrite-like arborizations, with more than one primary dendrite, extension of usually only one thin and long (0.5–1.6 mm) axon-like process and collaterals directed preferentially towards other Purkinje cells, developed with time in culture until the final form was reached by the equivalent of approximately day 16. Cell body size was 18–19 μm in diameter at this stage. Cell shapes were reminiscent of those described in certain cerebellar mouse mutants and in experimentally produced agranular cerebella. Many ultrastructural features of these cells correlated with those described for the in vivo counterpart. However, there was a lack of spiny branchlets and abnormally long persisting somatic spines. Synaptic contacts of the ‘en passant’ type could be seen at the Purkinje cell soma. Gray type I synapses were seen on Purkinje cell dendrites and spines.

Differentiation features of human pancreatic tumor cells maintained in nude mice and in culture: Immunocytochemical and ultrastructural studies

A poorly differentiated human pancreatic adenocarcinoma was maintained in nude mice for more than 3 years. When tumor fragments from xenografts were cultivated in suspension, some became adherent, allowing cell culture. Cytochemical, immunocytological and ultrastructural methods were used to study cell differentiation in both solid tumors and cultures. Pancreatic differentiation features such as cell polarization, production of secretory granules, and M1 and M3 mucus-associated antigens were maintained in the tumor cells, in vivo and in vitro. Moreover, in long-term cell cultures, cells were able to organize themselves spontaneously into duct-like structures. Other differentiation features such as production of pancreatic enzymes and hormones were not expressed. However, differentiation patterns such as an intestinal-like brush border and the presence of the M3 antigen associated with intestinal mucus were observed in both xenografts and cultures. This study shows the possible differentiation patterns which can be expressed by the hypothetical tumor pancreatic stem cell in nude mice as well as in culture.

Relationship between immune system and gram-negative bacteria. II. Natural killer cytotoxicity of Salmonella minnesota Rb 345-unbound human peripheral blood lymphocytes.

Spontaneous binding of human peripheral blood lymphocytes (PBL) to bacteria represents a promising approach for the characterization of lymphocyte subsets mediating different functions. In the light of previous findings on the high degree of spontaneous adherence of S. minnesota Rb cells to PBL, we have evaluated the natural killer (NK) cytotoxicity of PBL subpopulations that fail to bind to Rb bacteria. The S. minnesota Rb-unbound cell fraction exhibits higher levels of cytotoxic capacity, which is related to a more elevated frequency of active NK cells, as determined in an agarose-single cell cytotoxic assay. Moreover, the cytotoxic activity of the unbound fraction is additionally boosted by interferon-alpha pretreatment. The effector cells bear Fc gamma receptors that are involved in NK cell lysis, because a decrease of NK activity is observed after immune complex modulation of the receptors. Finally, these cells, which display a high percentage (approximately 70%) of typical large granular lymphocyte morphology, express HNK-1, T10, T8, and M1 antigens, and to a lesser extent T3 and T4 antigens. These data indicate a selective enrichment of NK cells in the S. minnesota Rb-unbound fraction.

Glial cells in the pineal gland of mice and rats. A combined immunofluorescence and electron-microscopic study.

Antigenic markers characteristic of astrocytes and their differentiative states (i.e., glial fibrillary acidic protein (GFAP), vimentin, and M1 and C1 antigens) were investigated in the pineal gland of mouse and rat using double immunolabeling techniques. In both species the so-called interstitial cells as characterized by TEM were shown to be astrocytes, since they expressed vimentin, but neither fibronectin (a marker for fibroblasts and endothelial cells) nor the neuron-specific L1 antigen or tetanus toxin receptors. Subpopulations of vimentin-positive pineal astrocytes were also GFAP- and C1- antigen-positive. M1- antigen-positive cells were not detected. It is concluded that a considerable proportion of interstitial cells in the pineal gland of rat and mouse are immature astrocytes which, in contrast to other parts of the central nervous system, persist into adulthood.

Colonic and gastric mucus-associated antigens: A comparative immunohistological study in precancerous and cancerous rat intestinal mucosa

Gastric M1 antigens were previously shown to be oncofetal markers for the colon in man and in the rat. They were observed very early during carcinogenesis in goblet cells of precancerous colonic mucosa; using an immunohistological method, we found that M1 were produced in 68% ( 41 60 ) of colonic adenocarcinomas and in 33% ( 21 63 ) of duodenal adenocarcinomas. M3C antigen has been described as being associated with human colonic mucus; in the rat it is restricted to the proximal colon. We found that M3C was produced in 91% ( 55 60 ) of colonic adenocarcinomas and in 15% ( 8 53 ) of duodenal adenocarcinomas. Before tumor appearance, M3C was sometimes expressed by goblet cells of the distal colon. We could not find it during fetal life. We have concluded that mucus-associated antigens can characterize modifications in cell differentiation in rat colonic carcinomas.

Expression of glial antigens C1 and M1 in the peripheral nervous system during development and regeneration

The expression of C1 and M1 antigens was studied by indirect immunofluorescence methods in histological sections of peripheral nerves and ganglia of C57BL/6J mice during development and regeneration. In sciatic nerves of adult mice, C1 but not M1 antigen is found in vimentin- and glial fibrillary acidic protein (GFAP)-positive Schwann cells. A similar distribution is also seen in trigeminal nerve, dorsal root and superior cervical ganglia, and olfactory nerve. In all cases vimentin-positive structures outnumber GFAP- or C1 antigen-positive ones. At birth, C1 antigen and vimentin are expressed in sciatic nerves, but GFAP is not yet detectable. M1 antigen cannot be detected in Schwann cells. In monolayer cultures of neonatal mouse dorsal root ganglia, C1 antigen is expressed in a fibrillary staining pattern in some, but not all morphologically identified Schwann cells. In vitro, M1 antigen is not detectable in Schwann cells. After lesioning sciatic nerves of adult mice by cut or crush, detectable levels of C1 antigen rise after 4–6 days: The number of immunofluorescently labeled structures and their relative intensities are drastically augmented, first distally more so than proximally, over control values from non-lesioned, i.e. contralateral nerves. A similar augmentation is also observed for vimentin and GFAP. M1 antigen expression does not reach detectable levels in Schwann cells under these conditions. The increased detectability of C1 antigen persists up to 150 days after lesioning, the longest time period tested.

Chick myotendinous antigen. II. A novel extracellular glycoprotein complex consisting of large disulfide-linked subunits.

This report describes the biochemical characterization of a novel extracellular matrix component, " myotendinous antigen," which appears early in chick limb morphogenesis at sites connecting developing muscle fibers, tendons, and bone ( Chiquet , M., and D. Fambrough , 1984; J. Cell Biol., 98:1926-1936). This extracellular matrix antigen is a major component of the secretory proteins released into the medium by fibroblast and muscle cultures; the soluble form is characterized here. This form of myotendinous antigen is a large glycoprotein complex consisting of several disulfide linked subunits (Mr approximately 150,000-240,000). The differently sized antigen subunits are related, since they yielded very similar proteolytic cleavage patterns. M1 antibody can bind to the denatured subunits. The antigen subunits, as well as a Mr approximately 80,000 pepsin-resistant antigenic domain derived from them, are resistant to bacterial collagenase. Despite possessing subunits similar in size to fibronectin, myotendinous antigen appears to be both structurally and antigenically unrelated to fibronectin or to other known extracellular matrix components. About seven times more M1 antigen per cell nucleus was released into the medium in fibroblast as compared to muscle cultures. In muscle conditioned medium, myotendinous antigen is noncovalently complexed to very high molecular weight material that could be heavily labeled by [3H]glucosamine and [35S]sulfate. This material is sensitive to chondroitinase ABC and hence appears to contain sulfated glycosaminoglycans. We speculate that myotendinous antigen might interact with proteoglycans on the surface of muscle fibers, thereby acting as a link to tendons.