Modified cerium-based and Gomori-based cerium methodes for light microscopic phosphatase histochemistry : The cerium-perhydroxide diaminobenzidine-nickel (Ce-H2O2-DAB-Ni and Ce/Ce-H2O2-DAB-Ni) two-step procedures

Kj, Halbhuber; Feuerstein, H; Möller, Ulrich; Klinger, Matthias

doi:10.1016/s0065-1281(11)80145-5

Cited by 7 publications

(9 citation statements)

References 45 publications

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“…For visual control of the reaction pattern in the conventional light microscope, the incubated and non-reoxidized sections (Gossrau et al 1989) were treated in H2Oa-containing DAB solution consisting of 0.015% DAB, 2% NiSO 4, 50 mM NaN 3, 0.005% H202 in 0.1 M acetate buffer for 60-120 min at 37 ~ (pH 6.0) or at 20 ~ (pH 6.5) (Halbhuber et al 1992), rinsed in ddH20, dehydrated over graded alcohol and coverslipped in Canada balsam.…”

Section: D-amino Acid Oxidase (Aaox) O;-hydroxy Acid Oxidase (Haox)mentioning

confidence: 99%

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Reflectance enzyme histochemistry (REH): visualization of cerium-based and DAB primary reaction products of phosphatases and oxidases in cryostat sections by confocal laser scanning microscopy

Scheven

Jirikowski

et al. 1996

Histochem Cell Biol

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In the present study the reflectance mode of confocal laser scanning microscopy was adapted to detect and to assess semiquantitatively cerium-based primary reaction products of oxidases [Ce(IV) perhydroxide] and phosphatases [Ce(III) hydroxyphosphate converted into Ce(IV) perhydroxyphosphate] as well as of the 3,3'-diaminobenzidine (DAB)-based primary reaction product of cytochrome c oxidase in cryostat sections. Confocal laser scanning microscopy offers a unique way of making visible histochemical reaction products which are weakly absorbant but sufficiently reflective. It was easily possible to record simultaneously the reflectance signals at the wavelength of the exciting laser (preferentially 488 nm) and the autofluorescence signals ( > 580 nm in our set-up) of glutaraldehyde-fixed tissue. The results of an imbibition study of cerium-containing model precipitates indicate that the cerium, generally, should be oxidized prior to observation because the index of refraction of Ce(IV) compounds is considerably higher than that of the corresponding Ce(III) compounds. An attempt at comparative numerical assessment of reflection intensities from reflectant parts in morphologically similar sections is presented. The proposed technique may open new possibilities in enzyme- and immunohistochemistry.

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Section: D-amino Acid Oxidase (Aaox) O;-hydroxy Acid Oxidase (Haox)mentioning

confidence: 99%

“…DAB and DAB-Ni (slightly modified DAB-Ni solution according to Halbhuber et al 1992; containing 0.015% DAB,. 2% NiSO 4, 2% dextrane T70, 0.005%-0.02% H202 in 0.1 M PIPES buffer, pH 6.5, at 20 ~ for 30-60 min were used to stain peroxidase precipitates.…”

Section: In Situ Hybridizationmentioning

confidence: 99%

Reflectance enzyme histochemistry (REH): visualization of cerium-based and DAB primary reaction products of phosphatases and oxidases in cryostat sections by confocal laser scanning microscopy

Scheven

Jirikowski

et al. 1996

Histochem Cell Biol

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“…Visualization of alkaline phosphatase activity via the conventional cerium phosphate procedure (Halbhuber et al 1992;von Noorden and Frederiks 1993) yields such a precipitate. Visualization of alkaline phosphatase activity via the conventional cerium phosphate procedure (Halbhuber et al 1992;von Noorden and Frederiks 1993) yields such a precipitate.…”

Section: Resultsmentioning

confidence: 99%

“…Visualization of alkaline phosphatase with the bromochloroindolyl phosphate/nitro blue tetrazolium (BCIP/NBT) method (McGadey 1970) or the conventional CPP technique (Halbhuber et al 1992), and peroxidase (Behringer et al 1991) with DAB were carried out according to published protocols.…”

Section: Immunoenzymatic Visualization Reactions For Light Microscopymentioning

confidence: 99%

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Specific visualization of precipitated cerium by energy-filtered transmission electron microscopy for detection of alkaline phosphatase in immunoenzymatic double labeling of tyrosine hydroxylase and serotonin in the rat olfactory bulb

Radcke

Stroh

Dworkowski

et al. 2002

Histochem Cell Biol

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To understand in detail the functional morphology of neuronal circuits it is important to identify at the ultrastructural level the incoming axon, its target neuron, and members of the signaling cascades involved. This, however, represents a formidable task, requiring highly sophisticated electron microscopic multiplelabeling techniques. To extend available double-labeling procedures such as combinations of immunogold and peroxidase methods, an additional, gold-and peroxidaseindependent procedure would represent a considerable advantage. The present investigation therefore aimed to use alkaline phosphatase as the immunoenzymatic label at the electron microscopic level via cerium phosphate precipitates. To our surprise we found that available techniques, which are well established for the visualization of endogenous enzymes in sections from various tissues, are not suitable for application to immunocytochemistry. Careful characterization of the individual reaction conditions, however, resulted in an optimized procedure with largely increased sensitivity. The novel technique yields cerium-containing precipitates which are massive enough to allow the detection of the immunoenzymatic reaction product in the electron microscope. Using the rat olfactory bulb as the model system we showed further that our technique allows the combination with the peroxidase/diaminobenzidine system for ultrastructural double labeling. For this purpose, the alkaline phosphatase product is identified by its cerium content via energy-filtered transmission electron microscopy and thereby differentiated from cerium-free peroxidase-derived precipitates. Doing so, we found that ascending serotoninergic fibers do not establish synapses with dopaminergic periglomerular cells in the rat olfactory bulb.

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Extralysosomal localisation of acid phosphatase in the rat kidney

Rüdiger

Kalicharan

et al. 1998

Histochemistry and Cell Biology

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There is strong evidence that acid phosphatase (AcPase) plays an important role in the catabolism of the glomerular basement membrane (GBM) and the removal of macromolecular debris resulting from ultrafiltration. Recent enzyme histochemical investigations provide new evidence of the antithrombotic and anti-inflammatory function of ADPase and on the distribution of AcPase in mouse kidney tubule cells. By means of 3 mM cerium as the trapping agent and 1 mM p-nitrophenyl phosphate as the substrate, extralysosomal AcPase could be demonstrated at the ultrastructural level. Following a mild perfusion fixation (2% formaldehyde + 0.07% glutaraldehyde), an effective postfixation and short enzyme incubations (20 min) with microwave irradiation, highly specific enzyme histochemical reaction product and reasonable structural preservation were obtained. Extralysosomal, membrane-bound AcPase was observed along the endoplasmic reticulum, the trans-Golgi cisternae, the nuclear envelope, basal infoldings of the proximal and distal tubular cells and on glomerular profiles, e.g. cell membranes of podocytes, endothelium and basement membrane. Large amounts of extralysosomal AcPase were observed in the basement membrane of glomeruli, in contrast to no AcPase activity in the tubular and mesangial basement membrane. The observed difference in AcPase activity in the tubular epithelial basement membrane and the GBM supports the idea that AcPase in GBM specifically serves in the clearance of macromolecular debris to facilitate ultrafiltration. In the GBM a laminar distribution is observed, suggesting that both epithelial and endothelial cells are involved in the production of AcPase.

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Modified cerium-based and Gomori-based cerium methodes for light microscopic phosphatase histochemistry : The cerium-perhydroxide diaminobenzidine-nickel (Ce-H2O2-DAB-Ni and Ce/Ce-H2O2-DAB-Ni) two-step procedures

Cited by 7 publications

References 45 publications

Reflectance enzyme histochemistry (REH): visualization of cerium-based and DAB primary reaction products of phosphatases and oxidases in cryostat sections by confocal laser scanning microscopy

Reflectance enzyme histochemistry (REH): visualization of cerium-based and DAB primary reaction products of phosphatases and oxidases in cryostat sections by confocal laser scanning microscopy

Specific visualization of precipitated cerium by energy-filtered transmission electron microscopy for detection of alkaline phosphatase in immunoenzymatic double labeling of tyrosine hydroxylase and serotonin in the rat olfactory bulb

Extralysosomal localisation of acid phosphatase in the rat kidney

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