Molecular hybridization of 3H-labelled ribosomal RNA with DNA in ultrathin sections prepared for electron microscopy

Jacob, J.; Todd, Katherine; Birnstiel, Max L.; Bird, Adrian

doi:10.1016/0005-2787(71)90746-5

Cited by 57 publications

(26 citation statements)

References 8 publications

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“…Early attempts to hybridize tritiated labelled 28S rRNA to rDNA of mouse L cells suggested that ribosomal cistrons were present in both intranucleolar and perinucleolar chromatin (Jacob et al, 1974). Since for a long time the in situ hybridization procedure could only be performed with radioactive probes and by means of autoradiography, the spatial resolution was necessarily limited.…”

Section: Rdna Localization By In Situ Hybridizationmentioning

confidence: 99%

Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

Thiry

Scheer

Goessens

1991

Electron Microscopy Reviews

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Abstract-Nucleoli are the morphological expression of the activity of a defined set of chromosomal segments bearing rRNA genes. The topological distribution and composition of the intranucleolar chromatin as well as the definition of nucleolar structures in which enzymes of the rDNA transcription machinery reside have been investigated in mammalian cells by various immunogold labelling approaches at the ultrastructural level. The precise intranucleolar location of rRNA genes has been further specified by electron microscopic in situ hybridization with a non-autoradiographic procedure.Our results indicate that the fibrillar centers are the sole nucleolar structures where rDNA, core histones, RNA polymerase I and DNA to po isomerase I are located together.Taking into account the potential value and limitations of immunoelectron microscopic techniques, we propose that transcription of the rRNA genes takes place within the confines of the fibrillar centers, probably close to the boundary regions to the surrounding dense fibrillar component.

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Section: Rdna Localization By In Situ Hybridizationmentioning

confidence: 99%

Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

Thiry

Scheer

Goessens

1991

Electron Microscopy Reviews

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“…Ultrastructural ISH for mRNA, which was first described by Jacob et al [5], has further been developed by several investigators [4, 6-9, 11-13, 18-25] and has been carried out in the three different approaches: preembedding method [4, 12-14, 20, 23, 25], ultrathin frozen sections [9,18,19], postembedding method [6-9, 12, 14]. In recent years, we have developed a non-radioisotopic EM-ISH method using biotinylated synthesized oligonucleotide probes for the ultrastructural visualization of growth hormone (GH) and prolactin (PRL) mRNAs in rat pituitary cells, and have applied this method to pathophysiological studies of pituitary cells [12][13][14].…”

Section: Introductionmentioning

confidence: 99%

Application of Ultrastructural In Situ Hybridization Combined with Immunohistochemistry to Pathophysiological Studies of Pituitary Cell: Technical Review.

Matsuno

Nagashima

Osamura

1998

Acta Histochem. Cytochem

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“…For this purpose, ISH methodologies have been developed that allow visualization of RNAs at the electron microscopic (EM) level. The first EM ISH report by Jacob et al (1) described a post-embedding method with radioactive labeled probes. The poor localization property of this method was fundamentally overcome by Binder et al (2) by applying a non-radioactive ISH method.…”

Section: Introductionmentioning

confidence: 99%

Saponin pre-treatment in pre-embedding electron microscopic in situ hybridization for detection of specific RNA sequences in cultured cells: a methodological study.

Macville

Wiesmeijer²,

Dirks³

et al. 1995

J Histochem Cytochem.

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We describe a method for detection of specific RNA targets in cultured cells at the electron microscopic (EM) level using pre-embedding in situ hybridization (ISH). The specimens were monitored by reflection-contrast microscopy (RCM) before processing for EM. A good balance between preservation of ultrastructure and intensity of hybridization signals was obtained by using mild aldehyde furation followed by saponin permeabilization. Digoxigenin-labeled probes were used for detection of human elongation factor (HEF) mRNA in HeLa cells, immediate early (E) mRNA in rat 9G cells, and 28s rRNA in both cell lines. The hybrids were detected immunocytochemically by the peroxidaseldiaminobenzidhe (DAB) method or by ultra-small gold with silver enhancement. Comparison of these methods favored the peroxi-

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Molecular hybridization of 3H-labelled ribosomal RNA with DNA in ultrathin sections prepared for electron microscopy

Cited by 57 publications

References 8 publications

Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

Localization of nucleolar chromatin by immunocytochemistry and in situ hybridization at the electron microscopic level

Application of Ultrastructural In Situ Hybridization Combined with Immunohistochemistry to Pathophysiological Studies of Pituitary Cell: Technical Review.

Saponin pre-treatment in pre-embedding electron microscopic in situ hybridization for detection of specific RNA sequences in cultured cells: a methodological study.

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