Visualization of the anionic sites in the cell wall of apple fruit using a cationic colloidal gold probe

1993 ◽  
Vol 51 ◽  
pp. 320-321
Author(s):  
Stéphane Roy ◽  
William S. Conway ◽  
Alley E. Watada ◽  
Christopher D. Pooley ◽  
William P. Wergin
Keyword(s):  
Cell Wall ◽  
Colloidal Gold ◽  
Apple Fruit ◽  
Gold Complex ◽  
Anionic Sites ◽  
Wall Strength ◽  
Anionic Binding Sites ◽  
Cationic Colloidal Gold ◽  
Softening Process ◽  
Gold Probe

The ripening of fleshy fruits involves a softening process that consists of biochemical changes in the cell wall and leads to cell separation. Calcium is an important constituent of the cell wall and plays roles in maintaining the firmness of fruit and in reducing postharvest decay. The modification of cell wall strength is believed to be influenced by calcium that interacts with acidic pectic polymers to form crossbridges. This study examined how the frequency and distribution of anionic binding sites in the cell walls of apple fruit were influenced by calcium infiltration.Mature “Golden Delicious” apple fruits were pressure infiltrated with either H2O or a 4% solution of CaCl2 and the pericarp was sampled and processed according to standard procedures. Cationic poly-Llysine colloidal gold complex was used in a one-step procedure to visualize anionic sites in muro. Observations were performed with light microscopy, following silver intensification, and with transmission electron microscopy.

Distribution of the anionic sites in the cell wall of apple fruit after calcium treatment

PROTOPLASMA ◽  
1994 ◽  
Vol 178 (3-4) ◽  
pp. 156-167 ◽  
Author(s):  
S. Roy ◽  
W. S. Conway ◽  
A. E. Watada ◽  
C. E. Sams ◽  
C. D. Pooley ◽  
...  
Keyword(s):  
Cell Wall ◽  
Apple Fruit ◽  
Calcium Treatment ◽  
Anionic Sites

scholarly journals Demonstration of anionic sites in human eccrine and apocrine sweat glands in post-embedded ultra-thin sections with cationic colloidal gold: effect of enzyme digestion on these anionic sites.

1993 ◽  
Vol 41 (8) ◽  
pp. 1197-1207 ◽  
Author(s):  
K Saga ◽  
M Takahashi
Keyword(s):  
Colloidal Gold ◽  
Dermatan Sulfate ◽  
Sweat Glands ◽  
Secretory Cells ◽  
Anionic Sites ◽  
Chondroitinase Abc ◽  
Dark Cell ◽  
Luminal Membrane ◽  
Cationic Colloidal Gold ◽  
Eccrine Sweat Glands

We localized anionic sites ultrastructurally in human eccrine and apocrine sweat glands with a poly-L-lysine-gold complex (cationic colloidal gold). Anionic sites were labeled by incubating Lowicryl K4M-embedded sections on droplets of cationic colloidal gold. In eccrine sweat glands, colloidal gold particles were restricted to the basolateral membrane of the secretory cells at low pH, whereas the luminal membrane did not react with the gold particles. Chondroitinase ABC digested these anionic sites. This indicates that chondroitin sulfate and/or dermatan sulfate constitutes anionic sites in the basal labyrinth of eccrine sweat glands. In apocrine sweat glands, the luminal membrane of the secretory cells showed strong reaction at low pH, whereas the contraluminal membrane did not show any reaction. Neuraminidase completely digested these anionic sites, which indicated that the anionic charge of the apocrine lumen was due to sialic acid. Differences in distribution and susceptibility to enzymes of anionic sites in cell membranes between eccrine and apocrine sweat glands may reflect functional differences between these glands. Dark cell granules in eccrine secretory cells were negative for the anionic sites when sections were labeled without any pre-treatment. However, pre-incubation of the grids on EGTA or deionized water unmasked the anionic sites on the dark cell granules. The positive staining after EGTA treatment was greatly decreased by reincubation with CaCl2. These results suggested that Ca blocked anionic sites in dark cell granules. Exposed anionic sites were digested with chondroitinase ABC. This indicated that chondroitinase ABC and/or dermatan sulfate composed the anionic sites in dark cell granules.

scholarly journals Detection of glomerular anionic sites in post-embedded ultra-thin sections using cationic colloidal gold.

1991 ◽  
Vol 39 (7) ◽  
pp. 965-972 ◽  
Author(s):  
N P Goode ◽  
M Shires ◽  
D M Crellin ◽  
A M Davison
Keyword(s):  
Hyaluronic Acid ◽  
Chondroitin Sulfate ◽  
Colloidal Gold ◽  
Endothelial Glycocalyx ◽  
Computer Assisted ◽  
Nuclear Staining ◽  
Anionic Sites ◽  
Thin Sections ◽  
Cationic Colloidal Gold ◽  
Influence Of Ph

We detected glomerular anionic sites in fixed, LR Gold-embedded ultra-thin tissue sections using cationic colloidal gold. Manual and computer-assisted quantitation were compared, and the influence of pH and glycosaminoglycan-degrading enzymes on site expression was examined. Both quantitation methods produced similar results. Alteration of pH within a narrow range (pH 2.5-3.0) markedly affected the staining pattern. At pH 2.5, epithelial and endothelial glycocalyx and regular sites restricted to the lamina rara externa were stained. At pH 3.0 and above, glycocalyx was unstained but intracellular and nuclear staining was present; glomerular basement membrane (GBM) and mesangial matrix sites were abundant. After chondroitinase ABC or hyaluronidase digestion, GBM staining was eliminated at pH 2.0 and reduced at pH 7.0 (p less than 0.001), suggesting that degraded sites are associated with chondroitin sulfate or hyaluronic acid. By contrast, prolonged heparitinase I digestion was ineffective at either pH. Digestion of purified substrates revealed crossreactivity of heparitinase towards chondroitin sulfate and of chondroitinase towards hyaluronic acid. Since tissue sites were reduced by chondroitinase but not heparitinase, we suggest that degradation is due to hyaluronidase activity of chondroitinase and the anionic sites are associated with hyaluronic acid. However, the influence of pH indicates that lamina rara externa sites are structurally distinct from other GBM anionic sites.

scholarly journals ROLE OF CALCIUM IN REDUCING POSTHARVEST CELL WALL DEGRADATION IN `GOLDEN DELICIOUS' APPLE FRUIT

HortScience ◽  
1995 ◽  
Vol 30 (2) ◽  
pp. 191a-191 ◽  
Author(s):  
S. Roy ◽  
W.S. Conway ◽  
A.E. Watada ◽  
G. Gillen ◽  
W.P. Wergin
Keyword(s):  
Cell Wall ◽  
Cell Walls ◽  
Secondary Ion Mass Spectrometry ◽  
Calcium Content ◽  
Apple Fruit ◽  
Postharvest Treatment ◽  
Significant Enrichment ◽  
Anionic Binding Sites ◽  
Cross Bridges

Calcium is an important constituent of the cell wall and plays roles in maintaining firmness of fruit and reducing postharvest decay. The modification of the cell wall is believed to be influenced by calcium that interacts with acidic pectic polymers to form cross-bridges. Infiltrating apples with CaCl2 has been suggested as an effective postharvest treatment for increasing the calcium content. Three different methodologies were used to analyze the effects of calcium on the cell walls: 1) nickel staining of polygalacturonate on free-hand sections, 2) cationic gold labeling of anionic binding sites in the cell walls, and 3) analytical detection of calcium ions (40Ca, 44Ca) using a secondary ion mass spectrometry. The combination of these methods allowed us to directly visualize the cellular features associated with the infiltration of calcium. Treatment resulted in significant enrichment in the cell wall of the pericarp, transformed the acidic pectins in calcium pectates, and resulted in new calcium cross-bridges. Evidence now suggests that exogenously applied calcium affects the cell wall by enhancing its strength and reinforcing adhesion between neighbor cells; therefore, calcium infiltration delays fruit degradation.

The Application of Protein A-Gold Immunocytochemistry to Studies of Protein Secretion

1985 ◽  
Vol 43 ◽  
pp. 426-429
Author(s):  
George H. Herbener ◽  
Antonio Nanci ◽  
Moise Bendayan
Keyword(s):  
Tissue Section ◽  
Colloidal Gold ◽  
Unit Area ◽  
Protein A ◽  
Extracellular Proteins ◽  
Gold Complex ◽  
Second Step ◽  
Igg Antibodies ◽  
Tissue Sections ◽  
Antigenic Sites

Protein A-gold immunocytochemistry is a two-step, post-embedding labeling procedure which may be applied to tissue sections to localize intra- and extracellular proteins. The key requisite for immunocytochemistry is the availability of the appropriate antibody to react in an immune response with the antigenic sites on the protein of interest. During the second step, protein A-gold complex is reacted with the antibody. This is a non- specific reaction in that protein A will combine with most IgG antibodies. The ‘label’ visualized in the electron microscope is colloidal gold. Since labeling is restricted to the surface of the tissue section and since colloidal gold is particulate, labeling density, i.e., the number of gold particles per unit area of tissue section, may be quantitated with ease and accuracy.

scholarly journals Latency- and Defense-Related Ultrastructural Characteristics of Apple Fruit Tissues Infected with Botryosphaeria dothidea

2001 ◽  
Vol 91 (2) ◽  
pp. 165-172 ◽  
Author(s):  
Ki Woo Kim ◽  
Eun Woo Park ◽  
Young Ho Kim ◽  
Kyung-Ku Ahn ◽  
Pan Gi Kim ◽  
...  
Keyword(s):  
Cell Wall ◽  
Fungal Growth ◽  
Apple Fruit ◽  
White Rot ◽  
Ultrastructural Changes ◽  
Disease Development ◽  
Botryosphaeria Dothidea ◽  
Transmission Electron ◽  
Thin Electron ◽  
Apple Fruits

Apple fruit tissues infected with Botryosphaeria dothidea were examined by transmission electron microscopy using susceptible cv. Fuji and resistant cv. Jonathan. Immature (green) and mature (red) fruits of cv. Fuji with restricted or expanding lesions were also examined to reveal subcellular characteristics related with latent and restricted disease development. In infected susceptible mature fruits, cytoplasmic degeneration and organelle disruption commonly occurred, accompanying cell wall dissolution around invading hyphae. Cell wall dissolution around invading hyphae in subepidermis was rare in immature, red halo-symptomed cv. Fuji and resistant cv. Jonathan fruits. In infected immature fruits of cv. Fuji, presumably at the latent state of disease development, cellular degeneration was less severe, and invading hyphae contained prominent microbody-lipid globule complexes or the deposition of thin electron-dense outer layer around cell wall of intercellular hyphae. Both mature fruits with red halos and resistant apple fruits formed cell wall protuberances at the outside of cell walls. In addition, electron-dense extramural layers were formed in the resistant apple fruits. Aberrant hyphal structures such as intrahyphal hyphae were found only in resistant fruit tissues, indicating the physiologically altered fungal growth. These ultrastructural changes of host tissues and fungal hyphae may reflect the pathogenesis of apple white rot under varying conditions of apple fruits.

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