Electron microscopy, immunostaining, cytoskeleton visualization, in situ hybridization, and three-dimensional reconstruction of Xenopus oocytes

Biliński, S; Jaglarz, Mariusz K.; Dougherty, Matthew; Kloc, Małgorzata

doi:10.1016/j.ymeth.2009.12.003

Cited by 13 publications

(18 citation statements)

References 17 publications

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“…Semithin sections (0.7-1.0 μm) were stained with 1% methylene blue in 1% borax and examined with a Leica DMR (Leica, Wetzlar, Germany) microscope. Ultrathin sections (70-100 nm) were contrasted with uranyl acetate and lead citrate according to standard protocols [14]; the sections were then examined in a JEOL 100SX transmission electron microscope (JEM, Japan) at an accelerating voltage of 80 kV.…”

Section: Embedding and Sectioningmentioning

confidence: 99%

Premature ovarian failure in nobox-deficient mice is caused by defects in somatic cell invasion and germ cell cyst breakdown

Lechowska

Biliński

Choi

et al. 2011

J Assist Reprod Genet

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Purpose To understand the mechanism of premature ovarian failure (POF). Methods The ultrastructural (electron microscopy) analysis of primordial ovarian follicles in Nobox deficient mice. Results We studied, for the first time, the fate of oogonia in embryonic (prenatal) mouse ovaries and showed that the abolishment of the transition from germ cell cysts to primordial follicles in the ovaries of Nobox deficient mice is caused by defects in germ cell cyst breakdown, leading to the formation of syncytial follicles instead of primordial follicles. Conclusions These results indicate that POF syndrome in Nobox deficient mice results from the faulty signaling between somatic and germ line components during embryonic development. In addition, the extremely unusual and abnormal presence of adherens junctions between unseparated oocytes within syncytial follicles indicates that faulty communication between somatic and germ cells is involved in, or leads to, abnormalities in the cell adhesion program.

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Section: Embedding and Sectioningmentioning

confidence: 99%

Premature ovarian failure in nobox-deficient mice is caused by defects in somatic cell invasion and germ cell cyst breakdown

Lechowska

Biliński

Choi

et al. 2011

J Assist Reprod Genet

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“…Histological images analyzed via specialist software can also be used for 3D reconstruction of internal organ structure. This method was applied in the reconstruction of the structure of testis cords within developing mouse testes (Nel-Themaat et al 2009) and ovarian follicles in Xenopus (Bilinski et al 2010).…”

Section: Histology: Light and Electron Microscopymentioning

confidence: 99%

“…Colloid gold or nanogold conjugated to secondary antibody are used in this technique. Both the pre-embedding and post-embedding versions of the IGS method were presented in detail by Bilinski et al (2010). In most commonly used post-embedding IGS methods, the tissue is gently fixed in PFA with or without glutaraldehyde (strong fixation and osmium tetroxide postfixation should be omitted in order not to disrupt immunoreactivity), dehydrated, embedded in hydrophilic resin, and cut on formvar-carbon-coated nickel grids; the grids with ultrathin sections are applied onto drops of the following solutions: blocking solution (BSA), primary antibodies, rinsing, secondary antibodies, rinsing, and contrast staining with uranyl acetate and lead citrate.…”

Section: Histology: Light and Electron Microscopymentioning

confidence: 99%

Methods for the Study of Gonadal Development

Piprek

2016

Results and Problems in Cell Differentiation

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Current knowledge on gonadal development and sex determination is the product of many decades of research involving a variety of scientific methods from different biological disciplines such as histology, genetics, biochemistry, and molecular biology. The earliest embryological investigations, followed by the invention of microscopy and staining methods, were based on histological examinations. The most robust development of histological staining techniques occurred in the second half of the nineteenth century and resulted in structural descriptions of gonadogenesis. These first studies on gonadal development were conducted on domesticated animals; however, currently the mouse is the most extensively studied species. The next key point in the study of gonadogenesis was the advancement of methods allowing for the in vitro culture of fetal gonads. For instance, this led to the description of the origin of cell lines forming the gonads. Protein detection using antibodies and immunolabeling methods and the use of reporter genes were also invaluable for developmental studies, enabling the visualization of the formation of gonadal structure. Recently, genetic and molecular biology techniques, especially gene expression analysis, have revolutionized studies on gonadogenesis and have provided insight into the molecular mechanisms that govern this process. The successive invention of new methods is reflected in the progress of research on gonadal development.

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“…The blocking, antibody incubation and washing steps were performed exactly as described in Bilinski et al (2010). Light microscopy cytokeratin immunostaining was performed by using fluorescein isothiocyanate (FITC)-conjugated C11 anti-pan cytokeratin antibody (cat.…”

Section: Light and Electron Microscopy Cytokeratin Immunostainingmentioning

confidence: 99%

“…The oocyte vegetal cortexes were observed under a Nikon fluorescence microscope. Electron microscopy immunostaining was performed as described in detail in Bilinski et al (2010).…”

Section: Light and Electron Microscopy Cytokeratin Immunostainingmentioning

confidence: 99%

Structural messenger RNA contains cytokeratin polymerization and depolymerization signals

et al. 2011

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We have previously shown that VegT mRNA plays a structural (translation-independent) role in the organization of the cytokeratin cytoskeleton in Xenopus oocytes. The depletion of VegT mRNA causes the fragmentation of the cytokeratin network in the vegetal cortex of Xenopus oocytes. This effect can be rescued by the injection of synthetic VegT RNA into the oocyte. Here, we show that the structural function of VegT mRNA in Xenopus oocyte depends on its combinatory signals for the induction or facilitation and for the maintenance of the depolymerization vs. polymerization status of cytokeratin filaments and that the 300-nucleotide fragment of VegT RNA isolated from the context of the entire molecule induces and maintains the depolymerization of cytokeratin filaments when injected into Xenopus oocytes. A computational analysis of three homologous Xenopus VegT mRNAs has revealed the presence, within this 300-nucleotide region, of a conserved base-pairing (hairpin) configuration that might function in RNA/protein interactions.

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Electron microscopy, immunostaining, cytoskeleton visualization, in situ hybridization, and three-dimensional reconstruction of Xenopus oocytes

Cited by 13 publications

References 17 publications

Premature ovarian failure in nobox-deficient mice is caused by defects in somatic cell invasion and germ cell cyst breakdown

Premature ovarian failure in nobox-deficient mice is caused by defects in somatic cell invasion and germ cell cyst breakdown

Methods for the Study of Gonadal Development

Structural messenger RNA contains cytokeratin polymerization and depolymerization signals

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