Visualization of Peroxisomes (Microbodies) and Mitochondria With Diaminobenzidine

Novikoff, Alex B.; Goldfischer, Sidney

doi:10.1177/17.10.675

Cited by 497 publications

(146 citation statements)

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“…Peroxidatic activity was demonstrated in microbodies of castor bean endosperm by the same procedures used to label liver microbodies (8,23). In castor bean endosperm, reaction product is distributed throughout the microbody matrix and, therefore, is not a unique characteristic of the crystalline or amorphous regions of the microbodies.…”

Section: Discussionmentioning

confidence: 99%

“…The tissues were either postfixed (see below) or allowed to warm to room temperature in the last buffer rinse and then incubated in one of the media listed below. These procedures are variations of several histochemical methods (8,11,23) Following incubation, the tissues were rinsed in four 10-min changes of buffer at 24 C and then postfixed in buffered OS04 for about 18 hr at 2 to 4 C. After postfixation, the tissues were rinsed in several changes of buffer and then dehydrated in a graded series of acetones and embedded in an Epon-Araldite epoxy resin mixture as described previously (21). Sections were cut with a Sorvall MT-1 microtome and viewed with a Philips microscope.…”

Section: Methodsmentioning

confidence: 99%

“…The tissues were either postfixed (see below) or allowed to warm to room temperature in the last buffer rinse and then incubated in one of the media listed below. These procedures are variations of several histochemical methods (8,11,23) (2,7,14,16). Both are utilized by the embryonic axis during germination and are largely gone by the 6th day of germination ( Fig.…”

mentioning

confidence: 99%

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Studies on Seeds

Mollenhauer

Totten

1970

Plant Physiol.

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Glyoxysomes, a form of microbody, are present in castor bean endosperm during the first 8 days of seed germination.They have a "typical" microbody form and are shown histochemically to contain catalase. The catalase label is distributed throughout the microbody and is not an exclusive feature of the crystalline or amorphous core.Castor bean endosperm contain a second cytosome, only slightly larger than the glyoxysomes, which is bound by a rough-surfaced membrane and which does not label for catalase. We have not observed these cytosomes in other tissues, suggesting that they may have a specific cellular function characteristic of castor bean endosperm.Glyoxysomes are a morphologically and biochemically unique component of many cells (3,4,12,13). They are a form of microbody containing enzymes of the glyoxylate cycle. Like peroxisomes and other types of microbodies, they also contain catalase and glycolic acid oxidase (4).Microbodies are present in castor bean endosperm throughout seed development, dormancy, and germination, but enzymes of the glyoxylate cycle do not reach maximal activity until the 4th or 5th day of seed germination (5, 12). There is no indication from ultrastructural studies that microbodies disappear, reform, or change significantly in number, in relation to the appearance of enzymes of the glyoxylate cycle. It is assumed, therefore, that glyoxysomes are somehow evolved from microbodies by enzyme activation or by gradual replacement of existing microbodies with those of the glyoxysome type.We have isolated microbodies, as well as glyoxysomes, from a number of seeds and have found that the glyoxysome fractions from castor bean endosperm sometimes contain a contaminating particle that is similar in size to glyoxysomes. Studies in vivo of germinating castor bean confirm that a microbody-like particle does exist in the endosperm and is most easily recognized when enzymes of the glyoxylate cycle are active. We thought initially that these new particles might be related, or equivalent, to glyoxysomes and, therefore, initiated these studies. This report describes these new particles and compares them structurally and histochemically with microbodies. 2"Studies on Seeds, I through IV" will appear in the Journal of Cell Biology. MATERIALS AND METHODSCastor bean (Hale variety and Baker variety 296 obtained from The Baker Castor Oil Co., La Mesa, Calif.) seeds were scarified and the seeds were soaked in water for 2 to 24 hr. The seeds were germinated for 3 to 8 days in the dark at 28 C in vermiculite moistened with distilled water.Slices of endosperm were prefixed in a mixture of 2% glutaraldehyde-2 % paraformaldehyde in 0.05 M collidine buffer containing 0.06 M sucrose, pH 7.3 to 7.4 for 2 hr, cold. After prefixation, the tissues were rinsed in six 5-to 10-min changes of buffer. Unless otherwise indicated, the buffer used was 0.05 M collidine with 0.06 M sucrose, pH 7.3 to 7.4. The tissues were either postfixed (see below) or allowed to warm to room temperature in the last buffer rinse and then ...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…The tissues were either postfixed (see below) or allowed to warm to room temperature in the last buffer rinse and then incubated in one of the media listed below. These procedures are variations of several histochemical methods (8,11,23) (2,7,14,16). Both are utilized by the embryonic axis during germination and are largely gone by the 6th day of germination ( Fig.…”

mentioning

confidence: 99%

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Studies on Seeds

Mollenhauer

Totten

1970

Plant Physiol.

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“…Cytochemical staining for peroxisomal catalase was performed following the method described by Novikoff and Goldfischer [9]. Small pieces of liver from each mouse were fixed with 3% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.4) for 1 h at 4°C, cut into 100-µm sections using an Oxford Vibratome (Oxford Laboratories, Foster City, CA, USA) and post-fixed with 3% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.4) for 1 h. The sections were incubated in a 3,3'-diaminobenzidine (DAB) reaction medium (0.2% DAB tetrahydrochloride, 50 mM propanediol, pH 9.7, 5 mM KCN, 0.05% H 2 O 2 ) for 1 h at room temperature, then post-fixed with aqueous 1% osmium tetroxide in 0.1 M sodium cacodylate buffer, dehydrated with a graded series of ethanol and acetone, and embedded in Epok 812 (Oken, Tokyo, Japan).…”

Section: Cytochemical Staining Of Peroxisomesmentioning

confidence: 99%

Peroxisome proliferator-activated receptor α-independent peroxisome proliferation

Zhang

Tanaka

Nakajima

et al. 2006

Biochemical and Biophysical Research Communications

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“…The liver of all animals was fixed in 10% neutral formalin, embedded in paraffin, sectioned at 3 µm, and stained with hematoxylin and eosin. For peroxisome visualization in the control and DHEA + PB groups, 45-µm-thick tissue slices fixed in 2.5% glutaraldehyde were reacted for 60 min at 37°C in alkaline 3,3'-diaminobenzidine medium [20] and postfixed in 1% OsO 4 . They were then embedded in epoxy resin, and 1 µm semi-thin sections were stained with toluidine blue.…”

mentioning

confidence: 99%

Prevention of Orotic-Acid-Induced Fatty Liver in Male Rats by Dehydroepiandrosterone and/or Phenobarbital.

Goto

Yamashita

Makita

1998

The Journal of Veterinary Medical Science

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ABSTRACT. Dehydroepiandrosterone (DHEA) is a steroid hormone which induces the peroxisome proliferation in rodents. The fatty liver induced by orotic acid and a high sucrose diet in male rats was prevented by the administration of DHEA and/or phenobarbital (PB). A significant increase in the liver weight was induced in the DHEA group (relative weight) and the DHEA + PB group (absolute and relative weight). The liver weight increased more conspicuously in the DHEA + PB group than in the DHEA group. The increase in the liver weight was caused by an increase in the cell size and peroxisome number. In addition, the administration of DHEA alone and the combination of DHEA and PB prevented the lipid droplet accumulation in hepatocytes. The administration of PB alone also prevented the accumulation of lipid droplets without any increase in the liver weight. -KEY WORDS: dehydroepiandrosterone, fatty liver, orotic acid, peroxisome proliferator, phenobarbital.J. Vet. Med. Sci. 60(4): 513-517, 1998 to 60%, a ventilation frequency of 13 to 15 air exchanges/ hr, and a 12-hr light/12-hr dark cycle (lights on: 07:00 to 19:00 hr). Control animals were fed the standard food for 2 weeks, and DHEA group rats were fed the standard food with 0.5% DHEA (Sigma Chemical Co.). Animals in the PB group were fed the standard food and intraperitoneally injected with 6% PB (Wako Pure Chemicals Co.) at 60 mg/ kg, once a day, 5 days a week, for 2 weeks. The DHEA + PB group were fed standard food with concomitant treatment with DHEA and PB. Body weight and food consumption were measured every 7 days. The final body weight was measured on the day of necropsy to calculate the relative organ weight. All animals were euthanized by exsanguination from abdominal veins and arteries following ether anesthesia and laparotomy. They were then examined for gross findings, and the absolute of the liver and organto-body weight ratio (relative weight) were determined. The liver of all animals was fixed in 10% neutral formalin, embedded in paraffin, sectioned at 3 µm, and stained with hematoxylin and eosin. For peroxisome visualization in the control and DHEA + PB groups, 45-µm-thick tissue slices fixed in 2.5% glutaraldehyde were reacted for 60 min at 37°C in alkaline 3,3'-diaminobenzidine medium [20] and postfixed in 1% OsO 4 . They were then embedded in epoxy resin, and 1 µm semi-thin sections were stained with toluidine blue. Statistical analysis: Body weight gain, food consumption, and organ weight were analyzed by one-way analysis of variance. When one-way analysis yielded a significant difference (P<0.05), the values were analyzed by Dunnett's test. RESULTSIn comparison with the control group, significant Dehydroepiandrosterone (3β-hydroxy-5-androsten-17-one, DHEA) is a metabolic intermediate in the testosterone, estrone, and estradiol synthesis pathway mainly in the human adrenal cortex and rodent gonads [1,28]. The actions of this hormone reported thus far included a possible antiobesity effect [2,16], preventive action on the development of athero...

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Visualization of Peroxisomes (Microbodies) and Mitochondria With Diaminobenzidine

Cited by 497 publications

References 0 publications

Studies on Seeds

Studies on Seeds

Peroxisome proliferator-activated receptor α-independent peroxisome proliferation

Prevention of Orotic-Acid-Induced Fatty Liver in Male Rats by Dehydroepiandrosterone and/or Phenobarbital.

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