scholarly journals Microfibril-associated glycoprotein-1 (MAGP-1) is specifically located on the beads of the beaded-filament structure for fibrillin-containing microfibrils as visualized by the rotary shadowing technique.

1996 ◽  
Vol 44 (12) ◽  
pp. 1389-1397 ◽  
Author(s):  
M Henderson ◽  
R Polewski ◽  
J C Fanning ◽  
M A Gibson
Keyword(s):  
Monoclonal Antibody ◽  
Immunoelectron Microscopy ◽  
Narrow Band ◽  
Immunogold Labeling ◽  
Structural Role ◽  
Filament Structure ◽  
Fibrillin 1 ◽  
Postembedding Immunoelectron Microscopy ◽  
Microscopic Techniques ◽  
Rotary Shadowing

This study used immunoelectron microscopic techniques to define the ultrastructural location of MAGP-1 on the fibrillin-containing microfibrils of the ocular zonule. A specific anti-MAGP-1 monoclonal antibody (MAb), 11B, was produced that did not crossreact with fibrillin-1 or other microfibrillar proteins. MAb 11B was shown by immunofluorescence to localize intensely to zonular tissue. Postembedding immunoelectron microscopy showed that MAGP-1 was associated with microfibrils throughout the zonule, with the exception of a narrow band of microfibrils at the junction with the lens capsule. With preembedding labeling, the anti-MAGP-1 MAb was found to localize in a crossbanding pattern, at intervals of about 50 nm, to microfibrils throughout the zonule and along bundles of microfibrils in surrounding vitreous tissue. Rotary shadowing of isolated microfibrils showed a "beads on a string" morphology with a periodicity of about 50 nm. With immunogold labeling, the anti-MAGP-1 antibody specifically localized on the beads in a symmetrical manner. Occasionally two gold partides were attached to the same bead, suggesting that multiple MAGP-1 molecules were present in the structure. The results indicate that MAGP-1 is intimately and regularly associated with the bead regions of fibrillin-containing microfibrils. The findings are consistent with a major structural role for MAGP-1 in microfibril biology.

Banded aggregates containing fibrillin are present in the cartilage matrix adjacent to chondrocytes in individuals affected with scoliosis

1992 ◽  
Vol 50 (1) ◽  
pp. 892-893
Author(s):  
Douglas R. Keene ◽  
Magaret Fairhurst ◽  
Catherine C. Ridgway ◽  
Lynn Y. Sakai
Keyword(s):  
Connective Tissue ◽  
Marfan Syndrome ◽  
Cross Section ◽  
Immunoelectron Microscopy ◽  
Cartilage Matrix ◽  
Negative Stain ◽  
Gene Coding ◽  
Rotary Shadowing

Matrix microfibrils are present in the connective tissue matrices of all tissues. Following standard TEM processing, they appear in cross section as cylindrical fibrils 8-10 nm in diameter, often associated with amorphous elastin. They are also seen in the absence of amorphous elastin, for example in the shallow papillary layer of skin, and also in cartilage matrix (Figure 1). Negative stain and rotary shadowing studies suggest that microfibrils are composed of laterally associated globular structures connected by fine filamentous strands (“ beaded strings”), and that they are extendable. Immunoelectron microscopy has demonstrated that fibrillin, a 350 Kd glycoprotein, is distributed along all microfibrils with a relaxed periodicity of about 54 nm The gene coding for fibrillin has recently been identified and is defective in the Marfan syndrome.

scholarly journals Monoclonal antibodies against chicken type IV and V collagens: electron microscopic mapping of the epitopes after rotary shadowing.

1984 ◽  
Vol 98 (5) ◽  
pp. 1637-1644 ◽  
Author(s):  
R Mayne ◽  
H Wiedemann ◽  
M H Irwin ◽  
R D Sanderson ◽  
J M Fitch ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Cleavage Site ◽  
Type Iv Collagen ◽  
Collagen Molecule ◽  
Electron Microscopic ◽  
Type V Collagen ◽  
Type Iv ◽  
Type V ◽  
Rotary Shadowing

The location of the epitopes for monoclonal antibodies against chicken type IV and type V collagens were directly determined in the electron microscope after rotary shadowing of antibody/collagen mixtures. Three monoclonal antibodies against type IV collagen were examined, each one of which was previously demonstrated to be specific for only one of the three pepsin-resistant fragments of the molecule. The three native fragments were designated (F1)2F2, F3, and 7S, and the antibodies that specifically recognize each fragment were called, respectively, IA8 , IIB12 , and ID2 . By electron microscopy, monoclonal antibody IA8 recognized an epitope located in the center of fragment (F1)2F2 and in tetramers of type IV collagen at a distance of 288 nm from the 7S domain, the region of overlap of four type IV molecules. Monoclonal antibody IIB12 , in contrast, recognized an epitope located only 73 nm from the 7S domain. This result therefore provides direct visual evidence that the F3 fragment is located closest to the 7S domain and the order of the fragments must be 7S-F3-(F1)2F2. The epitope for antibody ID2 was located in the overlap region of the 7S domain, and often several antibody molecules were observed to binding to a single 7S domain. The high frequency with which antibody molecules were observed to bind to fragments of type IV collagen suggests that there is a single population of type IV molecules of chain organization [alpha 1(IV)]2 alpha 2(IV), and that four identical molecules must form a tetramer that is joined in an antiparallel manner at the 7S domain. The monoclonal antibodies against type V collagen, called AB12 and DH2 , were both found to recognize epitopes close to one another, the epitopes being located 45-48 nm from one end of the type V collagen molecule. The significance of this result still remains uncertain, but suggests that this site is probably highly immunoreactive. It may also be related to the specific cleavage site of type V collagen by selected metalloproteinases and by alpha-thrombin. This cleavage site is also known to be located close to one end of the type V molecule.

The membrane skeleton of acetylcholine receptor domains in rat myotubes contains antiparallel homodimers of beta-spectrin in filaments quantitatively resembling those of erythrocytes

1995 ◽  
Vol 108 (9) ◽  
pp. 3145-3154 ◽  
Author(s):  
D.W. Pumplin
Keyword(s):  
Monoclonal Antibody ◽  
Acetylcholine Receptor ◽  
Acetylcholine Receptors ◽  
Cytoplasmic Membrane ◽  
Membrane Surface ◽  
Membrane Skeleton ◽  
Immunogold Labeling ◽  
Platinum Coating ◽  
High Concentrations ◽  
Receptor Clusters

I used immunogold labeling and quick-freeze, deep-etch, rotary replication to characterize the membrane skeleton at regions with high concentrations of acetylcholine receptor domains in receptor clusters of cultured rat muscle cells. This membrane skeleton consists of a network of filaments closely applied to the cytoplasmic membrane surface. The filaments are specifically decorated by immunogold labeling with a monoclonal antibody, VIIF7, that recognizes an isoform of beta-spectrin colocalizing with acetylcholine receptors. The filaments are 32 +/- 11 nm in length and three to four filaments (average 3.1-3.3) join at each intersection to form the network. These parameters are nearly identical to those reported previously for the membrane skeleton of erythrocytes. Depending on the amount of platinum coating, filament diameters range from 9 to 11 nm in diameter, and are 1.4 nm larger on average than spectrin filaments of erythrocytes replicated at the same time. Filaments are decorated with gold particles close to one end, consistent with the location of the epitope recognized by the monoclonal antibody. Computer modeling shows that all filament intersections in the membrane skeletal network are equally capable of being labeled by the monoclonal antibody. This pattern of labeling is consistent with a network containing antiparallel homodimers of beta-spectrin.

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