Pre-embedding Electron Microscopy Methods for Glycan Localization in Chemically Fixed Mammalian Tissue Using Horseradish Peroxidase-Conjugated Lectin

2010 ◽  
pp. 217-224 ◽  
Author(s):  
Yoshihiro Akimoto ◽  
Hayato Kawakami
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Mammalian Tissue

Comparative strengths and weaknesses of peroxidase and colloidal gold in pre- and post-embedding immunolabeling techniques

1987 ◽  
Vol 45 ◽  
pp. 1002-1005
Author(s):  
D. R. Abrahamson ◽  
P. L. St.John ◽  
E. W. Perry
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Light Microscopy ◽  
Colloidal Gold ◽  
Light Microscope ◽  
Ultrastructural Localization ◽  
The Other ◽  
Quantitative Studies ◽  
Dense Suspension ◽  
Antigen Localization

Antibodies coupled to tracers for electron microscopy have been instrumental in the ultrastructural localization of antigens within cells and tissues. Among the most popular tracers are horseradish peroxidase (HRP), an enzyme that yields an osmiophilic reaction product, and colloidal gold, an electron dense suspension of particles. Some advantages of IgG-HRP conjugates are that they are readily synthesized, relatively small, and the immunolabeling obtained in a given experiment can be evaluated in the light microscope. In contrast, colloidal gold conjugates are available in different size ranges and multiple labeling as well as quantitative studies can therefore be undertaken through particle counting. On the other hand, gold conjugates are generally larger than those of HRP but usually can not be visualized with light microscopy. Concern has been raised, however, that HRP reaction product, which is exquisitely sensitive when generated properly, may in some cases distribute to sites distant from the original binding of the conjugate and therefore result in spurious antigen localization.

Ultrastructural study of the avian ventral lateral geniculate nucleus

1991 ◽  
Vol 6 (2) ◽  
pp. 119-134 ◽  
Author(s):  
Gloria D. Guiloff
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Lateral Geniculate Nucleus ◽  
Ultrastructural Study ◽  
Lateral Geniculate

AbstractThe ultrastructure of the pigeon and quail ventral lateral geniculate nucleus was analyzed with standard electron microscopy and horseradish peroxidase tracing of its retinal and tectal afferents. Six types of neurons were distinguished: two large, two medium-sized, and two small types.

Pioneer neurones use basal lamina as a substratum for outgrowth in the embryonic grasshopper limb

Development ◽  
1988 ◽  
Vol 104 (4) ◽  
pp. 601-608 ◽  
Author(s):  
H. Anderson ◽  
R.P. Tucker
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Basal Lamina ◽  
Growth Cone ◽  
Growth Cones ◽  
Axon Outgrowth ◽  
Stages Of Development

During axonogenesis, contacts made by the growth cone with its substratum are important in guiding the direction of neurone outgrowth. This study examines the contacts made by the growth cones of pioneer neurones in the embryonic grasshopper limb. Individual pioneer neurones at different stages of development were injected with horseradish peroxidase and the contacts made by the filopodia at the tip of their growth cones were examined by electron microscopy. Filopodia made few contacts with mesodermal cells, some contacts with ectodermal cells and very frequent contacts with basal lamina underlying the ectoderm. Components of the basal lamina may therefore play a role in guiding pioneer axon outgrowth.

Fluorescent markers to study membrane retrieval in antidiuretic hormone-treated toad urinary bladder

1986 ◽  
Vol 251 (2) ◽  
pp. C274-C284 ◽  
Author(s):  
H. W. Harris ◽  
J. B. Wade ◽  
J. S. Handler
Keyword(s):  
Electron Microscopy ◽  
Urinary Bladder ◽  
Horseradish Peroxidase ◽  
Antidiuretic Hormone ◽  
Apical Membrane ◽  
Granular Cell ◽  
Cell Uptake ◽  
Toad Urinary Bladder ◽  
Osmotic Gradient ◽  
Apical Surface

Antidiuretic hormone (ADH) stimulation of toad urinary bladder causes fusion of intracellular vesicles called aggrephores with the apical plasma membrane of granular cells. Aggrephores contain intramembrane particle aggregates whose appearance in the apical membrane is believed to produce a large increase in its water permeability. ADH removal (ADH washout) is thought to cause the retrieval of aggrephores into granular cell cytoplasm. We studied granular cell uptake of dextran and horseradish peroxidase conjugated with fluorescein, rhodamine, or both during ADH washout. Granular cell uptake of fluorescent dextran was dependent on prior exposure to ADH, a linear function of dextran concentration, and increased by a transepithelial osmotic gradient. Immediately after removal of ADH, granular cell fluorescence was finely dispersed and located near the apical surface. Subsequently, it coalesced into larger bodies. This change was most apparent when a single bladder was subjected to two cycles of ADH stimulation and removal using a dextran containing a different fluorophore for each cycle. The ultrastructural correlate for these fluorescent patterns was identified using rhodamine-labeled horseradish peroxidase. Electron microscopy showed that after detachment from the apical membrane, label was initially in tubular-shaped vesicles near the apical surface. Later, these vesicles clustered near multivesicular bodies and transferred their label to these structures. These tubular vesicles closely resemble the morphology of aggrephores visualized by freeze-fracture electron microscopy. We conclude that these fluorescent compounds can be used as markers for the luminal contents of membrane retrieved during ADH washout and allow detailed study of its intracellular processing.

Horseradish peroxidase cDNA as a marker for electron microscopy in neurons

2007 ◽  
Vol 165 (2) ◽  
pp. 210-215 ◽  
Author(s):  
Thomas Schikorski ◽  
Samuel M. Young ◽  
Yunming Hu
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase

scholarly journals INTESTINAL CAPILLARIES

1969 ◽  
Vol 41 (1) ◽  
pp. 33-58 ◽  
Author(s):  
F. Clementi ◽  
G. E. Palade
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Intestinal Mucosa ◽  
Intercellular Junctions ◽  
Post Injection ◽  
Pore System ◽  
Blood Capillaries ◽  
Large Pore ◽  
Small Pore ◽  
Histochemical Procedure

Horseradish peroxidase (mol. diam. ≃50 A) and ferritin (mol. diam. ≃110 A) were used as probe molecules for the small and large pore system, respectively, in blood capillaries of the intestinal mucosa of the mouse. Peroxidase distribution was followed in time, after intravenous injection, by applying the Graham-Karnovsky histochemical procedure to aldehyde-fixed specimens. The tracer was found to leave the plasma rapidly and to reach the pericapillary spaces 1 min post injection. Between 1 min and 1 min 30 sec, gradients of peroxidase reaction product could be demonstrated regularly around the capillaries; their highs were located opposite the fenestrated parts of the endothelium. These gradients were replaced by even distribution past 1 min 30 sec. Ferritin, followed directly by electron microscopy, appeared in the pericapillary spaces 3–4 min after i.v. injection. Like peroxidase, it initially produced transient gradients with highs opposite the fenestrated parts of the endothelium. For both tracers, there was no evidence of movement through intercellular junctions, and transport by plasmalemmal vesicles appeared less efficient than outflow through fenestrae. It is concluded that, in the blood capillaries of the inintestinal mucosa, the diaphragms of the endothelial fenestrae contain the structural equivalents of the small pore system. The large pore system seems to be restricted to a fraction of the fenestral population which presumably consists of diaphragm-free or diaphragm-deficient units.

scholarly journals Immune complex receptors on cell surface. I. Ultrastructural demonstration of macrophages.

1976 ◽  
Vol 24 (8) ◽  
pp. 948-955 ◽  
Author(s):  
P E McKeever ◽  
A J Garvin ◽  
S S Spicer
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Cell Surface ◽  
Immune Complex ◽  
Immune Complexes ◽  
External Surface ◽  
Binding Capacity ◽  
Ultrastructural Localization ◽  
Soluble Complex ◽  
Receptor Sites

A method is described for ultrastructural localization of immune complex receptors on the surface of viable peritoneal exudate cells. The technique entails incubation with a soluble complex of horseradish peroxidase (HRP) and specific antibody to HRP at 4 degrees C followed by exposure to diaminobenzidine and processing for electron microscopy. The bound immune complexes were evident as focal deposits of HRP reaction product, adhering closely to the external surface of macrophages with an uninterrupted periodicity varying between 30 and 120 nm. Following incubation with an insoluble immune complex containing a higher proportion of antibody, receptor sites stained frequently, but large aggregates adhered to the cells. Rinsing cells after staining with soluble complexes partially displaced the bound immune complexes. Fixation prior to exposure to immune complexes largely eliminated the binding capacity of the immune complex receptors.

Sign in / Sign up

Export Citation Format

Share Document