scholarly journals ULTRASTRUCTURAL LOCALIZATION OF INTRACELLULAR IMMUNE GLOBULINS IN PLASMA CELLS AND LYMPHOBLASTS BY ENZYME-LABELED ANTIBODIES

1969 ◽  
Vol 17 (4) ◽  
pp. 211-224 ◽  
Author(s):  
ELIZABETH H. LEDUC ◽  
GEOFFREY B. SCOTT ◽  
STRATIS AVRAMEAS
Keyword(s):  
Immune Response ◽  
Plasma Cells ◽  
Ultrastructural Localization ◽  
Frozen Sections ◽  
Cytoplasmic Staining ◽  
Specific Antibodies ◽  
Perinuclear Space ◽  
Cacodylate Buffer ◽  
Immune Globulins ◽  
Ultrathin Frozen Sections

Techniques for localization of immune globulins by specific antibodies labeled with peroxidase or alkaline phosphatase were applied to spleens of rabbits hyperimmunized against various antigens. Best results were obtained with tissue blocks or thick sections after fixation in 1% formaldehyde (freshly prepared from paraformaldehyde) in cacodylate buffer with sucrose. The edges of blocks gave adequate penetration, comparable to thick sections. Ultrathin frozen sections which were reacted directly with labeled conjugate were less successful and glycol methacrylate sections failed to stain. Results were 3-fold: (1) immune globulins were demonstrated in ergastoplasmic cisternae, perinuclear space and Golgi apparatus of plasmocytes; (2) associated ribosomal staining varied somewhat with fixation and osmolarity of fixative; (3) lymphoblasts were shown to contain antibody. Generalized cytoplasmic staining was observed in the lymphoblasts, with no localization in the ergastoplasm and with no staining of control preparations. The significance of ribosomal and lymphoblast staining is discussed in relation to cell differentiation and the development of the immune response.

scholarly journals The ultrastructural localization of immunoglobulins in human b cells of immunoproliferative diseases

Blood ◽  
1982 ◽  
Vol 59 (6) ◽  
pp. 1132-1140 ◽  
Author(s):  
MF Gourdin ◽  
JP Farcet ◽  
F Reyes
Keyword(s):  
Endoplasmic Reticulum ◽  
B Cells ◽  
Plasma Cells ◽  
Simultaneous Detection ◽  
Ultrastructural Localization ◽  
Lymphocytic Leukemia ◽  
Cellular Distribution ◽  
Complex Plasma ◽  
Perinuclear Space ◽  
Human B Cells

Abstract The cellular distribution of immunoglobulins in human malignant and normal B cells was investigated by immunoelectron microscopy by direct incubation of fixed cells with electron microscopy by direct incubation of fixed cells with peroxidase-coupled antibody. These conjugates penetrated into the cell, resulting in the simultaneous detection of surface and cytoplasmic immunoglobulins. The latter were seen as specific intracisternal staining of the perinuclear space and endoplasmic reticulum and occasionally of the Golgi complex. Plasma cells were frequently characterized by a heterogeneity of reactivity of the endoplasmic reticulum. Minute amounts of cytoplasmic immunoglobulin were demonstrated in cells without developed secretory organelles, such as lymphoma cells and lymphocytes from chronic lymphocytic leukemia (CLL). The method allowed us to define several subsets of cells according to the expression of surface and cytoplasmic immunoglobulins and thus to determine the stage of maturation of cells involved in monoclonal proliferation.

scholarly journals The ultrastructural localization of immunoglobulins in human b cells of immunoproliferative diseases

Blood ◽  
1982 ◽  
Vol 59 (6) ◽  
pp. 1132-1140
Author(s):  
MF Gourdin ◽  
JP Farcet ◽  
F Reyes
Keyword(s):  
Endoplasmic Reticulum ◽  
B Cells ◽  
Plasma Cells ◽  
Simultaneous Detection ◽  
Ultrastructural Localization ◽  
Lymphocytic Leukemia ◽  
Cellular Distribution ◽  
Complex Plasma ◽  
Perinuclear Space ◽  
Human B Cells

The cellular distribution of immunoglobulins in human malignant and normal B cells was investigated by immunoelectron microscopy by direct incubation of fixed cells with electron microscopy by direct incubation of fixed cells with peroxidase-coupled antibody. These conjugates penetrated into the cell, resulting in the simultaneous detection of surface and cytoplasmic immunoglobulins. The latter were seen as specific intracisternal staining of the perinuclear space and endoplasmic reticulum and occasionally of the Golgi complex. Plasma cells were frequently characterized by a heterogeneity of reactivity of the endoplasmic reticulum. Minute amounts of cytoplasmic immunoglobulin were demonstrated in cells without developed secretory organelles, such as lymphoma cells and lymphocytes from chronic lymphocytic leukemia (CLL). The method allowed us to define several subsets of cells according to the expression of surface and cytoplasmic immunoglobulins and thus to determine the stage of maturation of cells involved in monoclonal proliferation.

scholarly journals ULTRASTRUCTURAL LOCALIZATION OF ANTIBODY IN DIFFERENTIATING PLASMA CELLS

1968 ◽  
Vol 127 (1) ◽  
pp. 109-118 ◽  
Author(s):  
Elizabeth H. Leduc ◽  
Stratis Avrameas ◽  
Michel Bouteille
Keyword(s):  
Electron Microscopy ◽  
Golgi Apparatus ◽  
Horseradish Peroxidase ◽  
Peroxidase Activity ◽  
Plasma Cells ◽  
Ultrastructural Localization ◽  
Fixed Tissue ◽  
Plasmacytic Differentiation ◽  
Perinuclear Space ◽  
Large Spherical

Antibody was localized by electron microscopy within differentiating and mature plasma cells of the spleens of hyperimmunized rabbits. Horseradish peroxidase was used as antigen. Intracellular antibody to peroxidase was revealed in glutaraldehyde-fixed tissue by coupling it with its antigen and then revealing the sites of peroxidase activity cytochemically. Antibody first appears in the perinuclear space of hemocytoblasts where it persists through differentiation into immature plasma cells, but it disappears from this site in mature plasma cells. Concomitant with the development of the ergastoplasm, antibody accumulates in many but not all of its cisternae. Antibody is present in the lamellar portion of the Golgi apparatus in all phases of plasmacytic differentiation. Mature plasma cells exhibit two types of antibody distribution, a concentration into large spherical intracisternal granules or an overflowing into all parts of the cytoplasm.

scholarly journals Presence of plasma cells binding autologous antibody during an immune response.

1979 ◽  
Vol 150 (5) ◽  
pp. 1265-1270 ◽  
Author(s):  
S Jackson ◽  
J Mestecky
Keyword(s):  
Immune Response ◽  
Human Serum Albumin ◽  
Serum Albumin ◽  
Plasma Cells ◽  
Fluorescein Isothiocyanate ◽  
Cross Reactivity ◽  
Lymphoid Tissues ◽  
Specific Antibodies ◽  
Tetramethylrhodamine Isothiocyanate ◽  
Idiotypic Antibody

Spleen and other lymphoid tissues of rabbits immunized with human serum albumin (HSA) and human lactoferrin (LF) were examined for the presence of cells forming anti-idiotype antibodies. To detect these cells, IgG, F(ab')2, or Fab' of specific antibodies were isolated, fluorochrome-tagged with tetramethylrhodamine isothiocyanate, and used as an idiotypic marker to detect splenic plasma cells that are producing anti-idiotypic antibody. By this procedure, we were able to demonstrate anti-idiotypic cells in surprisingly high numbers. For example, in six rabbits immunized with HSA for periods ranging from 36 to 542 d, the percentage of Ig-positive cells that stained with autologous idiotype ranged from 0.7 to 44; furthermore, cross-reactivity was observed among seven different anti-HSA preparations and two anti-LF antisera. The isotype of anti-idiotypic cells, determined by costaining with fluorescein isothiocyanate-labeled goat Fc-specific anti-rabbit Ig, was shown to be predominantly IgG. These findings provide evidence of the presence of plasma cells producing antibody to autologous idiotype during a vigorous immune response.

Immune Response of the Endolymphatic Sac to Horseradish Peroxidase: Immunologic Route from the Middle Ear to the Inner Ear

1989 ◽  
Vol 98 (12) ◽  
pp. 975-979 ◽  
Author(s):  
Minoru Ikeda ◽  
Claus Morgenstern
Keyword(s):  
Immune Response ◽  
Horseradish Peroxidase ◽  
Inner Ear ◽  
Middle Ear ◽  
Plasma Cells ◽  
Endolymphatic Sac ◽  
Local Immune Response ◽  
Frozen Sections ◽  
Response Region

Twenty guinea pigs were immunized with horseradish peroxidase (HRP) intradermally and challenged with 5 mg of the same antigen in the tympanic bulla. The appearance of immunoglobulin-producing cells (plasma cells) in the inner ear structure was examined immunohistochemically in frozen sections. Four to 10 days following antigen challenge, 5 of the 20 animals showed significantly increased plasma cells in the subepithelial connective tissue of the endolymphatic sac (ES). Those cells showed positive reactions, mainly with IgG followed by IgM. The cells that reacted positively with IgA were few. Some of these plasma cells were considered to contain the specific antibody against HRP. The results indicate the role of the ES as a local immune response region for the inner ear complex, as well as the existence of an immunologic route from the middle ear cavity to the inner ear, particularly to the ES.

scholarly journals Ultrastructural localization of the Ca2+ + Mg2+-dependent ATPase of sarcoplasmic reticulum in rat skeletal muscle by immunoferritin labeling of ultrathin frozen sections.

1982 ◽  
Vol 92 (2) ◽  
pp. 409-416 ◽  
Author(s):  
A O Jorgensen ◽  
A C Shen ◽  
D H MacLennan ◽  
K T Tokuyasu
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Plasma Membranes ◽  
Ultrastructural Localization ◽  
Published Data ◽  
Frozen Sections ◽  
Dependent Atpase ◽  
Sarcoplasmic Reticulum Membrane ◽  
Ultrathin Frozen Sections ◽  
Terminal Cisternae ◽  
Simultaneous Visualization

The ultrastructural localization of the Ca2+ + Mg2+-dependent ATPase of sarcoplasmic reticulum in rat gracilis muscle was determined by indirect immunoferritin labeling of ultrathin frozen sections. Simultaneous visualization of ferritin particles and of adsorption-stained cellular membranes showed that the Ca2+ + Mg2+-ATPase was concentrated in the longitudinal sarcoplasmic reticulum and in the nonjunctional regions of the terminal cisternae membrane but was virtually absent from mitochondria, plasma membranes, transverse tubules, and junctional sarcoplasmic reticulum. Ferritin particles were found preponderantly on the cytoplasmic surface of the membrane, in agreement with published data showing an asymmetry of the Ca2+ + Mg2+-ATPase within the sarcoplasmic reticulum membrane. Comparison of the density of ferritin particles in fast and slow myofibers suggested that the density of the Ca2+ + Mg2+-ATPase in the sarcoplasmic reticulum membrane in a fast myofiber is approximately two times higher than in a slow myofiber.

scholarly journals Ultrastructural localization of calsequestrin in rat skeletal muscle by immunoferritin labeling of ultrathin frozen sections.

1983 ◽  
Vol 97 (5) ◽  
pp. 1573-1581 ◽  
Author(s):  
A O Jorgensen ◽  
A C Shen ◽  
K P Campbell ◽  
D H MacLennan
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Central Region ◽  
Ultrastructural Localization ◽  
Frozen Sections ◽  
Rat Skeletal Muscle ◽  
Major Site ◽  
Ultrathin Frozen Sections ◽  
Terminal Cisternae

The ultrastructural localization of calsequestrin in rat skeletal muscle (gracilis) was determined by indirect immunoferritin labeling of ultrathin frozen sections. Calsequestrin was found in the lumen of transversely and longitudinally oriented terminal cisternae but was absent from most of the longitudinal sarcotubules and the fenestrated sarcoplasmic reticulum. Calsequestrin was occasionally observed in vesicular structures found in the central region of the I band. Since calsequestrin is believed to provide the major site of Ca2+ sequestration in the sarcoplasmic reticulum, the present results support the view that Ca2+, transported to the lumen of the sarcoplasmic reticulum, is preferentially sequestered in the terminal cisternae, but they also suggest that additional Ca2+ sequestration may occur near the center of the I band.

Immunoferritin localization of intracellular antigens in positively stained frozen sections

1978 ◽  
Vol 36 (2) ◽  
pp. 168-169
Author(s):  
K. T. Tokuyasu
Keyword(s):  
Surface Tension ◽  
Water Surface ◽  
Thin Layers ◽  
The Other ◽  
Positive Staining ◽  
Frozen Sections ◽  
The Past ◽  
Ultrathin Frozen Sections ◽  
Definition Of

During the past investigations of immunoferritin localization of intracellular antigens in ultrathin frozen sections, we found that the degree of negative staining required to delineate u1trastructural details was often too dense for the recognition of ferritin particles. The quality of positive staining of ultrathin frozen sections, on the other hand, has generally been far inferior to that attainable in conventional plastic embedded sections, particularly in the definition of membranes. As we discussed before, a main cause of this difficulty seemed to be the vulnerability of frozen sections to the damaging effects of air-water surface tension at the time of drying of the sections.Indeed, we found that the quality of positive staining is greatly improved when positively stained frozen sections are protected against the effects of surface tension by embedding them in thin layers of mechanically stable materials at the time of drying (unpublished).

Localization of pancreatic enzymes in ultrathin frozen sections stained with metal labeled antibody

1978 ◽  
Vol 36 (2) ◽  
pp. 90-91
Author(s):  
Kenjiro Yasuda
Keyword(s):  
Fluorescent Antibody ◽  
Pancreatic Enzymes ◽  
Tissue Preparation ◽  
Zymogen Granules ◽  
Frozen Sections ◽  
En Bloc ◽  
Antibody Method ◽  
Ultrathin Frozen Sections ◽  
Fluorescent Antibody Method

Localization of amylase,chymotrypsinogen and trypsinogen in pancreas was demonstrated by Yasuda and Coons (1966), by using fluorescent antibody method. These enzymes were naturally found in the zymogen granules. Among them, amylase showed a diffuse localization around the nucleus, in addition to the zymogen granules. Using ferritin antibody method, scattered ferritin granules were also found around the Golgi area (Yasuda et al.,1967). The recent advance in the tissue preparation enables the antigen to be localized in the ultrathin frozen sections, by applying the labeled antibodies onto the sections instead of staining the tissue en bloc.The present study deals with the comparison of the localization of amylase and lipase demonstrated by applying the bismuth-labeled, peroxidase-labeled and ferritin-labeled antibody methods on the ultrathin frozen sections of pancreas, and on the blocks of the same tissue.

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