Improvements in cytogenetic slide preparation: Controlled chromosome spreading, chemical aging and gradual denaturing

Henegariu, Octavian; Heerema, Nyla A.; Wright, Lisa Lowe; Bray‐Ward, Patricia; Ward, David C.; Vance, Gail H.

doi:10.1002/1097-0320(20010201)43:2<101::aid-cyto1024>3.0.co;2-8

Cited by 182 publications

(103 citation statements)

References 11 publications

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“…In contrast, Henegariu et al (2) specified no ambient conditions and used hot steam to moisten the slides and then put them on a 65-75°C metal plate, which, according to our own tests, dries the slides in 10 s. Whereas some researchers dropped cells on dry slides (4,5), others suggested that having a thin layer of water on the slides immediately before adding the cells improves spreading (1,2,6). Dropping cells from a height reportedly enhanced chromosome spreading (1,3), but others have disagreed (2,7). Aware of the "mysterious aspects" of chromosome spreading, Barch et al (10) suggested that it is more of an "art" than science.…”

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confidence: 81%

A new method for improving metaphase chromosome spreading

et al. 2002

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Background:The success of complex molecular cytogenetic studies depends on having properly spread chromosomes. However, inconsistency of optimum chromosome spreading remains a major problem in cytogenetic studies. Methods: The metaphase spreading process was carefully timed to identify the most critical phase of chromosome spreading. The effects of dropping height of cell suspension, slide condition, drying time, fixative ratio, and relative humidity on the quality of metaphase spreads were studied by quantitative examination of metaphase chromosome spreads. Normal and immortalized human epithelial ovarian cells, neuroblastoma cells, and normal lymphocytes were tested. Results: Humidity over the slide was the most important variable affecting the quality of chromosome spreads. Consistent improvement in chromosome spreading (larger metaphase area, less chromosome overlaps, or

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confidence: 81%

A new method for improving metaphase chromosome spreading

et al. 2002

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“…Fluorescent in situ hybridization was performed on freshly made slides 'aged' chemically as described by Henegariu et al (2001).…”

Section: Centromeric and Telomeric Signals In Nucleoplasmic Bridgesmentioning

confidence: 99%

Effects of hTERT on metal ion-induced genomic instability

et al. 2006

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There is currently a great interest in delayed chromosomal and other damaging effects of low-dose exposure to a variety of pollutants which appear collectively to act through induction of stress-response pathways related to oxidative stress and ageing. These have been studied mostly in the radiation field but evidence is accumulating that the mechanisms can also be triggered by chemicals, especially heavy metals. Humans are exposed to metals, including chromium (Cr) (VI) and vanadium (V) (V), from the environment, industry and surgical implants. Thus, the impact of low-dose stress responses may be larger than expected from individual toxicity projections. In this study, a short (24 h) exposure of human fibroblasts to low doses of Cr (VI) and V (V) caused both acute chromosome damage and genomic instability in the progeny of exposed cells for at least 30 days after exposure. Acutely, Cr (VI) caused chromatid breaks without aneuploidy while V (V) caused aneuploidy without chromatid breaks. The longerterm genomic instability was similar but depended on hTERT positivity. In telomerase-negative hTERTÀ cells, Cr (VI) and V (V) caused a long lasting and transmissible induction of dicentric chromosomes, nucleoplasmic bridges, micronuclei and aneuploidy. There was also a long term and transmissible reduction of clonogenic survival, with an increased b-galactosidase staining and apoptosis. This instability was not present in telomerasepositive hTERT þ cells. In contrast, in hTERT þ cells the metals caused a persistent induction of tetraploidy, which was not noted in hTERTÀ cells. The growth and survival of both metal-exposed hTERT þ and hTERTÀ cells differed if they were cultured at subconfluent levels or plated out as colonies. Genomic instability is considered to be a driving force towards cancer. This study suggests that the type of genomic instability in human cells may depend critically on whether they are telomerase-positive or -negative and that their sensitivities to metals could depend on whether they are clustered or diffuse.

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“…In brief, PBL metaphase and prometaphase chromosomes for FISH analysis were performed as described by Henegariu [7]. DNA probe preparation: Chromosome microdissection was performed according to previously described methods [8,9].…”

Section: Metaphase Chromosome Preparationsmentioning

confidence: 99%

Chromosome rearrangements in sublines of human embryonic stem cell lines hESM01 and hESM03

Карамышева¹,

Prokhorovich²,

Lagarkova³

et al. 2013

Biodiscovery

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Due to possible proliferative effects of karyotypic reorganization of human embryonic stem cell (hESC) lines detailed genetic analysis are indicated prior to any application of hESCs. Molecular cytogenetic analysis of two different hESC sublines was performed and revealed aberrant chromosomes in both of them, i.e. in hESM01r18 (46,ХХ,-18,+mar) and hESM0309 (46,ХХ,del(4),dup (9)). This study shows that microdissection and multicolor fl uorescence in situ hybridization (mFISH) can be used to detect the chromosomal changes precisely of the derivative chromosomes that are diffi cult to identify by conventional G-banded chromosome analysis. In the present study chromosome microdissection and reverse FISH were applied using multicolor fl uorescence in situ hybridization (mFISH) for detailed characterization of the derivative chromosomes. The karyotypes of hESC lines were described as: 46,ХХ,r(18)(::p11.31→q21.2::q21.2→p11.31::) and 46,XX,del(4)(q25q31.1),dup(9) (q12q33), respectively. The potential role of the chromosomal regions involved in rearrangements for cell proliferation is discussed.

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Improvements in cytogenetic slide preparation: Controlled chromosome spreading, chemical aging and gradual denaturing

Cited by 182 publications

References 11 publications

A new method for improving metaphase chromosome spreading

A new method for improving metaphase chromosome spreading

Effects of hTERT on metal ion-induced genomic instability

Chromosome rearrangements in sublines of human embryonic stem cell lines hESM01 and hESM03

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