Improvements in cytogenetic slide preparation

Controlled chromosome spreading, chemical aging and gradual denaturing

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

Research output: Contribution to journalArticle

149 Citations (Scopus)

Abstract

Background: Metaphase spreading is an essential technique for clinical and molecular cytogenetics. Results of classical banding techniques as well as complex fluorescent in situ hybridization (FISH) applications, such as comparative genomic hybridization (CGH) or multiplex FISH (M-FISH), are greatly influenced by the quality of chromosome spreading and pretreatment of the slide prior to hybridization. Materials and Methods: Using hot steam and a metal plate with a temperature gradient across its surface, a reproducible protocol for slide preparation, aging, and hybridization was developed. Results: This protocol yields good chromosome spreads from even the most difficult cell suspensions and is unaffected by the environmental conditions. Chromosome spreads were suitable for both banding and FISH techniques common to the cytogenetic laboratory. Chemical aging is a rapid slide pretreatment procedure for FISH applications, which allows freshly prepared cytogenetic slides to be used for in situ hybridization within 30 min, thus increasing analytical throughput and reducing bench-work. Furthermore, the gradually denaturing process described allows the use of fresh biologic material with optimal FISH results while protecting chromosomal integrity during denaturing. Conclusion: The slide preparation and slide pretreatment protocols can be performed in any laboratory, do not require specialized equipment, and provide robust results.

Original languageEnglish (US)
Pages (from-to)101-109
Number of pages9
JournalCytometry
Volume43
Issue number2
DOIs
StatePublished - Feb 1 2001
Externally publishedYes

Fingerprint

Fluorescence In Situ Hybridization
Cytogenetics
Chromosomes
Comparative Genomic Hybridization
Cytogenetic Analysis
Steam
Metaphase
In Situ Hybridization
Suspensions
Metals
Equipment and Supplies
Temperature

Keywords

  • Alcohol
  • CGH
  • Cytogenetics
  • FISH technique
  • G-banding
  • Hybridization
  • M-FISH
  • Metaphase spreading
  • Protocol
  • Slide aging

ASJC Scopus subject areas

  • Hematology
  • Cell Biology
  • Pathology and Forensic Medicine
  • Biophysics
  • Endocrinology

Cite this

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.

In: Cytometry, Vol. 43, No. 2, 01.02.2001, p. 101-109.

Research output: Contribution to journalArticle

Henegariu, Octavian ; Heerema, Nyla A. ; Wright, Lisa Lowe ; Bray-Ward, Patricia ; Ward, David C. ; Vance, Gail. / Improvements in cytogenetic slide preparation : Controlled chromosome spreading, chemical aging and gradual denaturing. In: Cytometry. 2001 ; Vol. 43, No. 2. pp. 101-109.
@article{7d055b46cfb84c9f83ab4d78ebc6e609,
title = "Improvements in cytogenetic slide preparation: Controlled chromosome spreading, chemical aging and gradual denaturing",
abstract = "Background: Metaphase spreading is an essential technique for clinical and molecular cytogenetics. Results of classical banding techniques as well as complex fluorescent in situ hybridization (FISH) applications, such as comparative genomic hybridization (CGH) or multiplex FISH (M-FISH), are greatly influenced by the quality of chromosome spreading and pretreatment of the slide prior to hybridization. Materials and Methods: Using hot steam and a metal plate with a temperature gradient across its surface, a reproducible protocol for slide preparation, aging, and hybridization was developed. Results: This protocol yields good chromosome spreads from even the most difficult cell suspensions and is unaffected by the environmental conditions. Chromosome spreads were suitable for both banding and FISH techniques common to the cytogenetic laboratory. Chemical aging is a rapid slide pretreatment procedure for FISH applications, which allows freshly prepared cytogenetic slides to be used for in situ hybridization within 30 min, thus increasing analytical throughput and reducing bench-work. Furthermore, the gradually denaturing process described allows the use of fresh biologic material with optimal FISH results while protecting chromosomal integrity during denaturing. Conclusion: The slide preparation and slide pretreatment protocols can be performed in any laboratory, do not require specialized equipment, and provide robust results.",
keywords = "Alcohol, CGH, Cytogenetics, FISH technique, G-banding, Hybridization, M-FISH, Metaphase spreading, Protocol, Slide aging",
author = "Octavian Henegariu and Heerema, {Nyla A.} and Wright, {Lisa Lowe} and Patricia Bray-Ward and Ward, {David C.} and Gail Vance",
year = "2001",
month = "2",
day = "1",
doi = "10.1002/1097-0320(20010201)43:2<101::AID-CYTO1024>3.0.CO;2-8",
language = "English (US)",
volume = "43",
pages = "101--109",
journal = "Cytometry",
issn = "0196-4763",
publisher = "John Wiley and Sons Inc.",
number = "2",

}

TY - JOUR

T1 - Improvements in cytogenetic slide preparation

T2 - Controlled chromosome spreading, chemical aging and gradual denaturing

AU - Henegariu, Octavian

AU - Heerema, Nyla A.

AU - Wright, Lisa Lowe

AU - Bray-Ward, Patricia

AU - Ward, David C.

AU - Vance, Gail

PY - 2001/2/1

Y1 - 2001/2/1

N2 - Background: Metaphase spreading is an essential technique for clinical and molecular cytogenetics. Results of classical banding techniques as well as complex fluorescent in situ hybridization (FISH) applications, such as comparative genomic hybridization (CGH) or multiplex FISH (M-FISH), are greatly influenced by the quality of chromosome spreading and pretreatment of the slide prior to hybridization. Materials and Methods: Using hot steam and a metal plate with a temperature gradient across its surface, a reproducible protocol for slide preparation, aging, and hybridization was developed. Results: This protocol yields good chromosome spreads from even the most difficult cell suspensions and is unaffected by the environmental conditions. Chromosome spreads were suitable for both banding and FISH techniques common to the cytogenetic laboratory. Chemical aging is a rapid slide pretreatment procedure for FISH applications, which allows freshly prepared cytogenetic slides to be used for in situ hybridization within 30 min, thus increasing analytical throughput and reducing bench-work. Furthermore, the gradually denaturing process described allows the use of fresh biologic material with optimal FISH results while protecting chromosomal integrity during denaturing. Conclusion: The slide preparation and slide pretreatment protocols can be performed in any laboratory, do not require specialized equipment, and provide robust results.

AB - Background: Metaphase spreading is an essential technique for clinical and molecular cytogenetics. Results of classical banding techniques as well as complex fluorescent in situ hybridization (FISH) applications, such as comparative genomic hybridization (CGH) or multiplex FISH (M-FISH), are greatly influenced by the quality of chromosome spreading and pretreatment of the slide prior to hybridization. Materials and Methods: Using hot steam and a metal plate with a temperature gradient across its surface, a reproducible protocol for slide preparation, aging, and hybridization was developed. Results: This protocol yields good chromosome spreads from even the most difficult cell suspensions and is unaffected by the environmental conditions. Chromosome spreads were suitable for both banding and FISH techniques common to the cytogenetic laboratory. Chemical aging is a rapid slide pretreatment procedure for FISH applications, which allows freshly prepared cytogenetic slides to be used for in situ hybridization within 30 min, thus increasing analytical throughput and reducing bench-work. Furthermore, the gradually denaturing process described allows the use of fresh biologic material with optimal FISH results while protecting chromosomal integrity during denaturing. Conclusion: The slide preparation and slide pretreatment protocols can be performed in any laboratory, do not require specialized equipment, and provide robust results.

KW - Alcohol

KW - CGH

KW - Cytogenetics

KW - FISH technique

KW - G-banding

KW - Hybridization

KW - M-FISH

KW - Metaphase spreading

KW - Protocol

KW - Slide aging

UR - http://www.scopus.com/inward/record.url?scp=0035241668&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0035241668&partnerID=8YFLogxK

U2 - 10.1002/1097-0320(20010201)43:2<101::AID-CYTO1024>3.0.CO;2-8

DO - 10.1002/1097-0320(20010201)43:2<101::AID-CYTO1024>3.0.CO;2-8

M3 - Article

VL - 43

SP - 101

EP - 109

JO - Cytometry

JF - Cytometry

SN - 0196-4763

IS - 2

ER -