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W B Stallcup

Publications and source records attributed to W B Stallcup.

At least 55 records · Page 3Linked to original sources
PubMed1990

Interaction of the NG2 chondroitin sulfate proteoglycan with type VI collagen.

The NG2 chondroitin sulfate proteoglycan is a membrane-associated molecule of approximately 500 kD with a core glycoprotein of 300 kD. Both the complete proteoglycan and a smaller quantity of the 300-kD core are immunoprecipitable with polyclonal and monoclonal antibodies against purified NG2. From some cell lines, the antibodies coprecipitate NG2 and type VI collagen, the latter appearing on SDS-PAGE as components of 140 and 250 kD under reducing conditions. The immunoprecipitation of type VI collagen does not seem to be due to recognition of the collagen by the antibodies, but rather to binding of the collagen to NG2. Studies on the NG2-type VI collagen complex suggest that binding between the two molecules is mediated by protein-protein interactions rather than by ionic interactions involving the glycosaminoglycans. Immunofluorescence double labeling in frozen sections of embryonic rat shows that NG2 and type VI collagen are colocalized in structures such as the intervertebral discs and arteries of the spinal column. In vitro the two molecules are highly colocalized on the surface of several cell lines. Treatment of these cells resulting in a change in the distribution of NG2 on the cell surface also causes a parallel change in type VI collagen distribution. Our results suggest that cell surface NG2 may mediate cellular interactions with the extracellular matrix by binding to type VI collagen.

Animals↗
PubMed1989

A neuroectoderm-associated ganglioside participates in fibronectin receptor-mediated adhesion of germinal cells to fibronectin.

Previous studies with a rat neural cell line have shown that the D1.1 ganglioside, an O-acetylated derivative of GD3, is involved in cellular adhesion to fibronectin. In vivo, D1.1 is present on germinal cells of the developing rat central nervous system, but not on postmitotic cells that migrate away from the germinal zones. These observations suggest that D1.1 could participate in adhesive interactions with germinal zones and that the loss of D1.1 could be involved in the decision to being migration. In support of this hypothesis, immunofluorescence histochemistry shows that both fibronectin and fibronectin receptor are colocalized with D1.1 in the ventricular zones of the embryonic rat brain and in the external granule cell layer of the postnatal cerebellum. Dishes coated with monoclonal antibody against D1.1 were used to isolate D1.1-positive germinal cells from Embryonic Day 14 cerebrum and from Postnatal Day 6 cerebellum. These cells are able to adhere to fibronectin-coated dishes by a mechanism that is inhibitable by a synthetic hexapeptide containing the arg-gly-asp cell recognition sequence of fibronectin. Adhesion is also partially inhibited by antibody against fibronectin receptor and is slowed by anti-D1.1 antibody, implicating both the receptor and the ganglioside in the adhesion process. During 3 days in culture these D1.1-positive, fibronectin receptor-positive cells exhibit a neuronal phenotype, as judged by morphology and staining with tetanus toxin. This further confirms the neuroepithelial origin of the cells. The cells do not synthesize detectable amounts of fibronectin, thus leaving unidentified the source of the fibronectin seen in the germinal zones in tissue sections. Immunoprecipitation experiments show that the fibronectin receptors present on these cells are heterodimers. Under nonreducing conditions, the immunoprecipitates contain an alpha-subunit of 150-160 kDa and a beta-subunit of 115-125 kDa.

Age Factors↗
PubMed1989

Characterization of a partial cDNA clone for the NILE glycoprotein and identification of the encoded polypeptide domain.

A partial cDNA clone [2.4 kilobase (kb)] for the nerve growth factor-inducible large external (NILE) glycoprotein was selected from a lambda gt 11 expression library constructed using mRNA from PC 12 cells. A 0.2 kb subclone (pNILE-1B) was used for Northern blot analysis of NILE message present in 2 NILE-positive neuronal cell lines and 2 NILE-negative glial cell lines. pNILE-1B hybridizes with components of 6.8 and 2.0 kb in the 2 neuronal cell lines but fails to show hybridization with any components in the 2 glial cell lines. Only the 6.8 kb species would be large enough to code for the NILE polypeptide. A rabbit antiserum was prepared against the NILE-beta-galactosidase fusion protein produced by the NILE clone. This antiserum (anti-NILE-beta-gal) immunoprecipitates NILE glycoprotein from neuronal cell lines, further confirming the authenticity of the NILE cDNA clone. The epitope recognized by anti-NILE-beta-gal is contained in an 85 kDa tryptic fragment from the phosphorylated carboxy terminus of NILE. The 160 kDa tryptic fragment containing the amino terminus is not recognized by anti-NILE-beta-gal. Both immunoprecipitation and immunofluorescence experiments indicate that the anti-NILE-beta-gal epitope is not exposed on the cell surface but is accessible only after cells are treated with detergent. The cytoplasmic nature of the determinant is also indicated by its absence on a truncated, soluble form of NILE released from cells (possibly by a proteolytic mechanism) into the medium.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies↗
PubMed1989

Characterization of a partial cDNA clone for the NILE glycoprotein and identification of the encoded polypeptide domain.

A partial cDNA clone [2.4 kilobase (kb)] for the nerve growth factor-inducible large external (NILE) glycoprotein was selected from a lambda gt11 expression library constructed using mRNA from PC 12 cells. A 0.2 kb subclone (pNILE-1B) was used for Northern blot analysis of NILE message present in 2 NILE-positive neuronal cell lines and 2 NILE-negative glial cell lines. pNILE-1B hybridizes with components of 6.8 and 2.0 kb in the 2 neuronal cell lines but fails to show hybridization with any components in the 2 glial cell lines. Only the 6.8 kb species would be large enough to code for the NILE polypeptide. A rabbit antiserum was prepared against the NILE-beta-galactosidase fusion protein produced by the NILE clone. This antiserum (anti-NILE-beta-gal) immunoprecipitates NILE glycoprotein from neuronal cell lines, further confirming the authenticity of the NILE cDNA clone. The epitope recognized by anti-NILE-beta-gal is contained in an 85 kDa tryptic fragment from the phosphorylated carboxy terminus of NILE. The 160 kDa tryptic fragment containing the amino terminus is not recognized by anti-NILE-beta-gal. Both immunoprecipitation and immunofluorescence experiments indicate that the anti-NILE-beta-gal epitope is not exposed on the cell surface but is accessible only after cells are treated with detergent. The cytoplasmic nature of the determinant is also indicated by its absence on a truncated, soluble form of NILE released from cells (possibly by a proteolytic mechanism) into the medium. This released NILE is 15-20 kDa smaller than the detergent-extracted NILE and, in addition to lacking the anti-NILE-beta-gal epitope, does not contain the cytoplasmic site(s) of phosphorylation. Nucleotide sequencing of the pNILE-1B subclone confirms the location of the anti-NILE-beta-gal epitope in the cytoplasmic domain. The clone contains an open reading frame coding for a 79 amino acid segment of the polypeptide that differs in only 2 residues from the cytoplasmic domain of the L1 glycoprotein.

Amino Acid Sequence↗
PubMed1988

Melanoma-associated gangliosides in the fish genus Xiphophorus.

Gangliosides from benign and malignant melanomas and from normal skin of the fish genus Xiphophorus were isolated and analyzed by thin-layer chromatography. Individual ganglioside components were characterized by mapping according to their sialic acid content and by cleavage with neuraminidases. In all three tissues examined, sulfatide and the gangliosides NeuAc-GalCer (GM4), II3NeuAc-LacCer (GM3), II3NeuAc-GgOse3Cer (GM2), and II3(NeuAc)2-LacCer (GD3) were found. Ganglioside GD3 yielded a positive reaction, following immunoadsorption with mouse monoclonal antibody R24 on thin-layer plates. Two alkali-labile disialoganglioside species were specifically recognized by mouse monoclonal antibody D1.1, thus indicating the presence of O-acetyl-neuraminic acid residues. One of them, a major ganglioside component of the malignant melanoma, was identified as O-acetyl-GD3, since it could be converted to the R24-positive GD3 ganglioside after alkaline saponification. The other one appears to be restricted to the malignant tumor and represents a novel melanoma-associated ganglioside derivative. It was characterized as O-acetyl(NeuAc)2-nLc4Cer by exoglycosidase cleavage, by proving its neutral carbohydrate backbone as type II-chain lacto-series oligosaccharide using mouse monoclonal antibody 1B2, and by its cross-reaction with antibody R24 following alkaline treatment. Using antibody R24 and cryopreserved tissue sections of both benign and malignant amelanotic melanomas from albino fishes, it was demonstrated that one of the main melanoma-associated gangliosides, GD3, was exposed predominantly in the malignant tumor. Thus, the chemical nature and even the immunohistochemical localization of the gangliosides in fish melanomas proved to be very similar to those of the known gangliosides in the phylogenetically distant human melanomas.

Animals↗
PubMed1988

Involvement of gangliosides and glycoprotein fibronectin receptors in cellular adhesion to fibronectin.

We have used a rat neural cell line, B65, to investigate the relative contributions of gangliosides and glycoprotein receptors in adhesion to fibronectin. Monoclonal antibodies against two neuroectoderm-associated gangliosides, D1.1 and GD3, inhibit the rate of B65 attachment to fibronectin, suggesting that these gangliosides are involved in the adhesion process. Adhesion to fibronectin is not affected by a third monoclonal antibody against a separate, unidentified cell-surface component of B65 cells. Furthermore, B65 cells lacking D1.1 adhere to fibronectin at a slower rate than B65 cells that express D1.1. The involvement of glycoprotein receptors in adhesion is demonstrated by the ability of antibodies against human fibronectin receptor to inhibit B65 attachment to fibronectin. In addition, adhesion is blocked by a hexapeptide containing the Arg-Gly-Asp fibronectin sequence which is necessary for binding to the receptor. Trypsin treatment of B65 cells in the absence of divalent cations results in proteolysis of the fibronectin receptor with an accompanying loss of ability of the cells to attach to fibronectin. D1.1 and GD3 expression is not affected by this trypsinization, indicating that the gangliosides alone are incapable of mediating attachment. The glycoprotein receptors must be primarily responsible for adhesion to fibronectin with the gangliosides playing a secondary role as enhancers or modulators.

Animals↗
PubMed1988

Altered expression of the D1.1 ganglioside in the cerebellum of the Weaver mouse.

Expression of the D1.1 ganglioside was studied immunohistochemically in developing cerebella from normal and weaver mutant mice. In the normal cerebellum at postnatal day 7 (P7), D1.1 expression was restricted to the external granule-cell layer (EGL). At later ages, D1.1 disappeared as the developing granule neurons ceased mitosis and began migrating toward the internal granule-cell layer. In the weaver cerebellum, D1.1 was expressed in the EGL in apparently normal fashion at P7, but failed to disappear at later ages. As late as P35, D1.1 immunoreactivity was observed throughout the weaver cerebellar cortex. The relative amounts of D1.1 ganglioside in weaver and normal cerebella were compared by thin layer chromatography of total gangliosides, followed by overlay of the chromatogram with anti-D1.1 and 125I-labelled second antibody. Autoradiograms showed that at P12 and P35 the weaver tissue contains six- to tenfold more D1.1 than normal tissue. These findings suggest that one result of the weaver mutation is prolonged expression of D1.1. We speculate that the D1.1 ganglioside might be involved in adhesive interactions that regulate the timing of granule-cell migration from the EGL. The prolonged expression of D1.1 could be responsible, in part, for the failure of granule-cell migration in the weaver cerebellum.

Aging↗
PubMed1987

Plasticity of developing cerebellar cells in vitro studied with antibodies against the NG2 antigen.

The NG2 antigen, a chondroitin-sulfate proteoglycan, is a cell surface marker for a class of smooth protoplasmic astrocytes found throughout the brain and at high frequency in the cerebellar molecular layer (Levine and Card, 1987). To study the development of the cerebellar astrocytes at the level of the single cell, we have analyzed the distribution of the NG2 antigen by indirect immunofluorescence in dissociated cell cultures prepared from postnatal cerebella and compared that distribution to the distribution of several other cell surface and intracellular antigens that identify specific cell types in cultures of nervous tissue. When cerebellar cells from 5 d rat pups were grown in a medium containing 10% fetal calf serum, the NG2-labeled cells, which constituted 0.1-1.0% of the total glial cells present, contained glial fibrillary acidic protein (GFAP)-immunoreactive filaments and bound monoclonal antibody A2B5, a surface marker for neurons and some astrocytes. Approximately 30% of the NG2-labeled cells were also labeled with tetanus toxin, an additional surface marker for neurons and immature astrocytes. Less than 2% of the cells were labeled with antibodies against galactocerebroside or with monoclonal antibody O1, both of which are surface markers for oligodendrocytes. About half the NG2-labeled cells exhibited high-affinity uptake of 3H-GABA, and this uptake was partially inhibited by both beta-alanine and DABA. Thus, the NG2 antigen is a cell surface marker for a subpopulation of the type II or fibrous astrocytes present in the cultures. When the cerebellar cells were grown in a chemically defined, serum-free medium, the NG2-labeled cells had a stellate morphology and between 50-60% of the cells bound tetanus toxin. Although almost all the cells bound antibody A2B5, less than 5% of the cells expressed either of the oligodendrocyte surface markers or GFAP immunoreactivity. As was the case with cells grown in serum-containing medium, 60% of the NG2-labeled cells had high-affinity uptake of 3H-GABA. However, this uptake was inhibited by DABA but not by beta-alanine. This phenotype may be the in vitro analog of the NG2-labeled, filament-lacking, smooth protoplasmic astrocytes identified in the intact adult cerebellum. The expression of these 2 phenotypes could be reversed by switching the tissue culture medium within 5 d of plating the cells. These results demonstrate that the in vitro environment can influence the phenotypic properties expressed by developing cerebellar astrocytes and suggest that smooth protoplasmic astrocytes may be developmentally related to glial cells of the O-2A lineage.

Animals↗
PubMed1987

Bipotential glial precursor cells of the optic nerve express the NG2 proteoglycan.

We have studied the expression of the NG2 chondroitin sulfate proteoglycan on bipotential glial precursor cells in cultures of postnatal rat optic nerve. Purified populations of these precursor cells were prepared by panning dissociated optic nerve cells on dishes coated with monoclonal A2B5 antibody. Using immunofluorescence double staining, we found that NG2 was present on almost 95% of the purified A2B5+ precursor cells. The NG2 core protein from optic nerve cells was identified by immune precipitation and PAGE and was found to be identical to the 300,000 Da NG2 core protein from a clonal rat cell line B49. Over a culture period of 5 d in medium containing 10% fetal calf serum, more than 80% of the NG2+ precursor cells acquired the glial fibrillary acidic protein (GFAP), an astrocyte-specific marker. Under these conditions, fewer than 10% of the NG2+ cells expressed galactocerebroside (GC), an oligodendrocyte-specific marker. These GFAP+GC- type II astrocytes continued to express the NG2 antigen for up to 10 d in culture. During a 5 d culture period in hormonally supplemented, serum-free medium, fewer than 15% of the NG2+ cells expressed GFAP, while up to 40% expressed GC. The NG2 antigen continued to be expressed for only a short period of time by these GFAP-GC+ oligodendrocytes, so that mature oligodendrocytes in the cultures became NG2-. These results support our previous suggestion that the NG2 antigen is found on a class of neural cells that can differentiate along more than one pathway.

Animals↗
PubMed1987

The nerve growth factor-inducible large external (NILE) glycoprotein and neural cell adhesion molecule (N-CAM) have distinct patterns of expression in the developing rat central nervous system.

The nerve growth factor-inducible large external (NILE) glycoprotein and the neural cell adhesion molecule (N-CAM) have both been implicated in the process of nerve fiber fasciculation. To evaluate the respective roles of the 2 molecules in fiber tract formation, we used immunohistochemical means to compare their distributions in the developing rat central nervous system. In the spinal cord, hindbrain, forebrain, retina, and cerebellum, N-CAM was present on undifferentiated cells in germinal zones as well as on differentiating cells and in nerve fiber tracts. In contrast, NILE was restricted to the developing fiber tracts in all these areas. No fiber tracts were found that were obviously lacking one or the other of the 2 molecules during the period of tract development. However, in all cases except that of the cerebellar molecular layer, nerve fiber tracts appeared to lose NILE and retain N-CAM after the major phases of tract development were completed. The fact that NILE is restricted to nerve fiber tracts during relatively short but crucial phases of tract development suggests that NILE plays a very specific role in the formation of fiber bundles. The more ubiquitous N-CAM molecule may have a more general role in neural histogenesis.

Animals↗
PubMed1986

Localization of a neurectoderm-associated cell surface antigen in the developing and adult rat.

The distribution of a neurectoderm-associated carbohydrate antigen (termed D1.1) in tissues of the developing and adult rat was determined using indirect immunofluorescent techniques. The antigen was detected as early as embryonic days 8 and 9 when it was localized to cells within the developing neural plate and neural tube. As the central nervous system (CNS) developed, the anti-D1.1 antibody labeled neuroepithelial cells but not terminally differentiated neurons or glial cells. In addition, the notochord and somatic mesoderm were labeled transiently with the antibody. Outside of the CNS, the antibody labeled dorsal root ganglia neurons, adrenal chromaffin cells and cells of the kidney glomerulus. These tissues were labeled at embryonic day 14 and the labeling persisted in the adult. We used a sensitive immunoautoradiography assay to identify antigenic gangliosides present in extracts of these tissues. The anti-D1.1 antibody recognized a ganglioside of kidney and adrenal glands that has a chromatographic mobility identical to that of the D1.1 antigen previously identified from cell lines and developing cerebellum. However, the antibody bound to a separate and distinct set of gangliosides present in extracts of adult dorsal root ganglia. Thus, the carbohydrate sequence recognized by the antibody can be associated with more than one molecular species of ganglioside. These results demonstrate that within the context of the developing CNS, the D1.1 antigen is a stage-specific embryonic antigen, but, as is the case with other cell surface carbohydrate antigens, is also found on a limited but developmentally unrelated set of tissues in the adult.

Animals↗
PubMed1985

Polymorphism among NILE-related glycoproteins from different types of neurons.

We have used biosynthetic, degradative and immunochemical techniques to examine some of the structural features of the NILE (nerve growth factor inducible large external) glycoprotein. Biosynthetic experiments show that the NILE species present on PC12 cells has a core polypeptide with a molecular weight of 160,000 dalton. This polypeptide is initially glycosylated by immature, high mannose oligosaccharides to yield a precursor glycoprotein of 190,000 dalton. Oligosaccharide processing of this precursor in the Golgi apparatus yields the mature 230,000 dalton NILE glycoprotein. NILE-related glycoproteins from different neuronal cell types have molecular weights ranging from 215,000 to 230,000 dalton. Degradative experiments using neuraminidase to remove sialic acid residues and endoglycosidase F to remove all N-linked oligosaccharides indicate that the polymorphism of the NILE-related glycoproteins may be due to differences both at the polypeptide level and at the level of glycosylation. Although NILE glycoproteins and the neural cell adhesion molecule (N-CAM) both appear to be involved in neurite fasciculation, NILE and N-CAM components can be shown to be different from each other by several criteria. (1) The electrophoretic mobilities of NILE and N-CAM are different. NILE-related molecules from various sources migrate as single bands in the range of 215,000-230,000 dalton, while N-CAM from these same sources can exist as 3 separate components of 180,000, 140,000 and 120,000 dalton. (2) Rabbit antibody against NILE does not recognize N-CAM, and rabbit antibody against N-CAM does not recognize NILE. (3) N-CAM is present on some cell types that lack NILE. (4) The core polypeptides of NILE components have apparent molecular weights of about 160,000 dalton, while the N-CAM core polypeptide is about 120,000 dalton in size.

Animals↗
PubMed1985

Involvement of the nerve growth factor-inducible large external glycoprotein (NILE) in neurite fasciculation in primary cultures of rat brain.

The nerve growth factor-inducible large external glycoprotein (NILE) has been found only on the surface of neuronal cells and Schwann cells. Since NILE seems to be concentrated on neurites, we have investigated its possible role in the development of neurites in primary cultures of rat brain. Cultures of embryonic day 14 (E14) whole brain and cultures of postnatal day 5 (P5) cerebellum were grown in the presence of Fab' fragments of antibody against NILE in an attempt to perturb the normal pattern of neurite development. For comparison, cultures were treated with two other reagents that recognize neuronal cell surface molecules: tetanus toxin, which binds to the GD1b and GT1 gangliosides, and Fab' fragments of antibody against neural cell adhesion molecule (N-CAM). Under the conditions used, none of the exogenous reagents affected neurite outgrowth, but specific effects on neurite fasciculation were observed. Anti-NILE inhibited fasciculation in cultures of E14 whole brain but had no effect on fasciculation in cultures of P5 cerebellum. Conversely, anti-N-CAM inhibited fasciculation in cultures of P5 cerebellum, which contain the adult form of N-CAM, but had little effect on fasciculation in cultures of E14 whole brain, which contain the embryonic form of N-CAM. Tetanus toxin had no effect on fasciculation in either culture system. Our results imply that NILE-mediated neurite-neurite interactions are stronger than N-CAM (embryonic)-mediated interactions in the E14 brain cultures, whereas N-CAM (adult)-mediated interactions are stronger than NILE-mediated interactions in the P5 cerebellar cultures.

Animals↗
PubMed1985

Antibody against nerve growth factor-inducible large external (NILE) glycoprotein labels nerve fiber tracts in the developing rat nervous system.

The NILE (nerve growth factor-inducible large external) glycoprotein is a 230,000-dalton molecule found on the surface of PC12 cells. Immunologically cross-reactive glycoproteins in the molecular weight range of 215,000 to 230,000 have been found on many types of neurons in culture. Using immunohistochemical methods, we have shown that NILE-related glycoproteins are present in neuronal fiber tracts of the developing rat brain. Antibody against the NILE glycoprotein specifically labels processes that appear identical to those recognized by antibodies against the neurofilament triplet of proteins. These processes are clearly distinct from the radial glial fibers recognized by antibody against the intermediate filament protein vimentin. NILE glycoprotein is not distributed uniformly over the entire neuronal surface but is concentrated on neurites and is much less abundant on cell bodies. NILE-positive fiber tracts are first seen in the spinal cord and rhombencephalon on embryonic day 11 and over the next 2 days appear in the mesencephalon and diencephalon. Staining in the telencephalon is not seen until embryonic day 15. The appearance of NILE immunoreactivity in these various regions closely parallels the appearance of neurofilament polypeptides, suggesting that NILE-related glycoproteins are present during the early phases of fiber tract formation. This idea is supported by the finding that the NILE glycoprotein can be found postnatally in parts of the nervous-system such as the cerebellar cortex and olfactory bulb which undergo major histogenesis during the postnatal period. In the cerebellum the appearance of NILE immunoreactivity in the two major fiber zones, the molecular layer and the white matter, parallels the development of the fiber structure of these layers. These findings support tissue culture studies which suggest a role for the NILE glycoprotein in mediating nerve fiber fasciculation.

Animals↗
PubMed1984

A monoclonal antibody recognizes an O-acylated sialic acid in a human melanoma-associated ganglioside.

Monoclonal antibody D1.1 originally prepared against the B49 cell line derived from a rat brain tumor was shown to react with a ganglioside present in fetal rat brain. We have found that this antigen is also present in human malignant melanoma tumors as well as many melanoma cell lines. The ganglioside from human melanoma cell lines migrates between GM1 and GM2 on one-dimensional thin layer chromatography. Analysis by two-dimensional thin layer chromatography with intermediate ammonia treatment suggests that the ganglioside contains one or more base-labile O-acyl esters. Mild base hydrolysis under conditions known to remove O-acyl esters results in complete loss of antigenic reactivity. Thus, the alkali-labile moiety is a critical component of the epitope recognized by the antibody. Analysis of the sialic acids of total gangliosides from [6-3H]glucosamine-labeled melanoma cells showed that approximately 10% of these molecules are O-acylated. Similar analysis of the purified ganglioside showed that greater than 30% of the sialic acids comigrated with authentic 9-O-acetyl-N-acetylneuraminic acid. The antibody did not cross-react with normal human skin melanocytes nor with any of a large number of normal human adult and fetal tissues. The antibody also did not react with numerous other malignant cell lines studied. These findings suggest that the antigenic epitope defined by antibody D1.1 contains an O-acylated sialic acid and may arise from aberrant O-acetylation occurring in human malignant melanoma cells.

Animals↗
PubMed1984

The D1.1 antigen: a cell surface marker for germinal cells of the central nervous system.

A monoclonal antibody against a cell surface ganglioside present on neuroepithelial cells was produced by immunizing mice with the B49 cell line, a clonal line with properties of both neurons and glial cells. The expression of this antigen, designated as D1.1, was analyzed in the developing rat cerebellum. The D1.1 antigen was localized by the immunofluorescent staining method to germinal cells of the external granule layer (EGL). Fluorescent labeling of cells comprising the EGL was apparent on embryonic day 18 when the EGL first forms, and the labeling was present throughout the period of postnatal cerebellar development. No cells within the adult cerebellum were labeled with the anti-D1.1 antibody. The D1.1-labeled cells of the EGL synthesize DNA, as demonstrated by [3H] thymidine autoradiography. However, within 48 hr after their final mitosis, nascent cerebellar cells that had migrated away from the external granule layer were no longer labeled with antibody. Some of the neurons and some of the astrocytes in cerebellar cell cultures were fluorescently labeled with the anti-D1.1 antibody. The number of anti-D1.1-labeled neurons in the cultures decreased over the first 10 days in vitro in agreement with the findings that in vivo the fluorescent labeling of the D1.1 antigen disappears from postmitotic cells. The antibody recognizes a ganglioside that in thin layer chromatographic experiments has a mobility between that of the GM1 and GM2 ganglioside. These data suggest that the D1.1 ganglioside antigen is a cell surface marker for germinal cells and that the acquisition and subsequent loss of this antigen is an aspect of the biochemical maturation of neurons and glial cells.

Animals↗