Fibroblasts of skin fragments as a tool for the investigation of genetic diseases: technical recommendations

Coelho, Janice Carneiro; Giugliani, Roberto

doi:10.1590/s1415-47572000000200004

Cited by 26 publications

(13 citation statements)

References 4 publications

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“…Upon total confluence, the fibroblasts were divided by addition of 0.25% trypsin-EDTA [Coelho and Giugliani, 2000]. When a sufficient number of cells was obtained, fibroblasts were promptly submitted to neuraminidase determination using the method of O'Brien and Warner [1980], which is based on enzyme cleavage of a synthetic substrate (4-methylumbelieferyl-a-N-acetylneuraminic acid) at 378C, pH 4.3, for 30 min.…”

Section: Methodsmentioning

confidence: 99%

Application of a protocol for the detection of disorders of sialic acid metabolism to 124 high‐risk Brazilian patients

Castilhos

Mello

Burin

et al. 2003

American J of Med Genetics Pt A

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Lysosomal storage disorders (LSD) present great clinical variability. Included in this group are sialic acid metabolism disorders (SAMD). In the present study, we describe the application of a 3-step protocol for the diagnosis of SAMD, including (1). oligosaccharide and sialyloligosaccharide chromatography; (2). quantitative determination of sialic acid; and (3). measurement of neuraminidase activity. Application of our protocol to 124 individuals at risk for SAMD led to the diagnosis of five affected patients, two with type I sialidosis, one with type II sialidosis, and two with galactosialidosis. Due to its simplicity and efficiency, we propose the use of this protocol for the diagnostic evaluation of patients with suspected SAMD, which could be specially useful to non-specialized laboratories and to services located in developing countries.

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Section: Methodsmentioning

confidence: 99%

Application of a protocol for the detection of disorders of sialic acid metabolism to 124 high‐risk Brazilian patients

Castilhos

Mello

Burin

et al. 2003

American J of Med Genetics Pt A

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“…Fibroblast cell culture The fibroblast cell culture was performed according to the method described by Coelho & Giugliani (2000) with a few modifications. Briefly, skin fragments from the back of the patient and her parents were aseptically removed with a punch and immediately transferred to a sterile flask containing Ham's F‐10 nutrient mixture (Life Technologies, Gaithersburg, MD, USA) supplemented with 30% FCS.…”

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confidence: 99%

β‐Spectrin S^ta Bárbara: a novel frameshift mutation in hereditary spherocytosis associated with detectable levels of mRNA and a germ cell line mosaicism

Bassères¹,

Duarte

Hassoun

et al. 2001

Br J Haematol

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Hereditary spherocytosis (HS) is a common inherited anaemia characterized by the presence of spherocytic red cells and by a heterogeneous nature in terms of its clinical presentation, molecular basis and inheritance. Defects in several membrane protein genes have been involved in the pathogenesis of HS, including defects in the beta-spectrin gene. We detected a novel frameshift mutation in the beta-spectrin gene, a C deletion at codon 638, in a patient presenting with HS and spectrin deficiency. The mutant protein was not detected in the membrane or in other cellular compartments, but detectable levels of mutant mRNA were found in the patient. Interestingly, this mutation was not present in the patient's parents, suggesting a genetic mosaicism, especially as the patient has an affected brother with the same molecular defect. We analysed DNA from different tissues of the parents and the mutation was absent from all tissues analysed. This mutation seems to be confined to the germ cell lineage of the patient's mother and must present a mosaic pattern in these cells as the patient also has unaffected siblings.

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“…The American embryologist Ross Granville Harrison (1870Harrison ( -1959 developed the first techniques of cell culture in vitro in the first decade of the twentieth century [52][53][54][55][56]. In Harrison's experiments (1907)(1908)(1909)(1910), at the Yale University), small pieces of living frog embryonic tissue were isolated and grew outside the body.…”

Section: Harrison's Hanging Drop Technique and Dr Carell's Immortal mentioning

confidence: 99%

History of Cell Culture

Jędrzejczak-Silicka¹

2017

New Insights Into Cell Culture Technology

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From the ancient Romans, through the Middle Ages, to the late of the nineteenth century, the Aristotelian doctrine of spontaneous generation was one of the most basic laws. Even the invention of the microscope and investigations of Leeuwenhoek and Hook did not disprove the Aritostelian doctrine. Finally, in the eighteenth century, the spontaneous generation doctrine was laid by Louis Pasteur. Moreover, in the first decade of the eighteenth century, nucleus was observed in plant and animal tissues, and Virchow and other scientists presented the view that cells are formed via scission of preexisting cells. In the first decade of the twentieth century, Ross Harrison developed the first techniques of cell culture in vitro, and Burrows and Carrel improved Harrison's cell cultures. In mid-twentieth century, the basic principles for plant and animal cell cultures in vitro were developed, and human diploid cell lines were established. On the basis of knowledge about the cell cycle and gene expression regulation, the first therapeutic proteins were produced using mammalian cell cultures. The end of twentieth century and early twenty-first century brought the progress in 3-D cell culture technology and created the possibility of the tissue engineering and the regenerative medicine development.

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Fibroblasts of skin fragments as a tool for the investigation of genetic diseases: technical recommendations

Cited by 26 publications

References 4 publications

Application of a protocol for the detection of disorders of sialic acid metabolism to 124 high‐risk Brazilian patients

Application of a protocol for the detection of disorders of sialic acid metabolism to 124 high‐risk Brazilian patients

β‐Spectrin S^ta Bárbara: a novel frameshift mutation in hereditary spherocytosis associated with detectable levels of mRNA and a germ cell line mosaicism

History of Cell Culture

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Fibroblasts of skin fragments as a tool for the investigation of genetic diseases: technical recommendations

Cited by 26 publications

References 4 publications

Application of a protocol for the detection of disorders of sialic acid metabolism to 124 high‐risk Brazilian patients

Application of a protocol for the detection of disorders of sialic acid metabolism to 124 high‐risk Brazilian patients

β‐Spectrin Sta Bárbara: a novel frameshift mutation in hereditary spherocytosis associated with detectable levels of mRNA and a germ cell line mosaicism

History of Cell Culture

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β‐Spectrin S^ta Bárbara: a novel frameshift mutation in hereditary spherocytosis associated with detectable levels of mRNA and a germ cell line mosaicism