Optimized fixation and storage conditions for FISH analysis of single- cell suspensions

Sarah Murrell-Bussell; Dianne Nguyen; Wendy D. Schober; Jeffrey Scott; Joe Leigh Simpson; Sherman Elias; Farideh Z. Bischoff; Dorothy E. Lewis

doi:10.1177/002215549804600811

Optimized fixation and storage conditions for FISH analysis of single- cell suspensions

Sarah Murrell-Bussell, Dianne Nguyen, Wendy D. Schober, Jeffrey Scott, Joe Leigh Simpson, Sherman Elias, Farideh Z. Bischoff, Dorothy E. Lewis

Research output: Contribution to journal › Article › peer-review

8 Scopus citations

Abstract

In our protocol to isolate and identify fetal cells in maternal peripheral blood, antibody (Ab)-stained cells are preserved with paraformaldehyde (PF) before batch flow cytometric sorting. However, PF fixation compromises the quality of subsequent interphase fluorescence in situ hybridization (FISH). We therefore examined the effect of PF concentrations and storage time in phosphate-buffered saline (PBS) on the quality of FISH signals. Cells were analyzed for changes in light scatter, morphology, and accessibility of target cell DNA. Fixation in 3% PF for 1 hr was ideal for both flow cytometry and subsequent FISH detection. However, beyond 10 days of storage, FISH quality deteriorated.

Original language	English (US)
Pages (from-to)	971-973
Number of pages	3
Journal	Journal of Histochemistry and Cytochemistry
Volume	46
Issue number	8
DOIs	https://doi.org/10.1177/002215549804600811
State	Published - Aug 1998

Keywords

Cell fixation methods
Flow cytometry
Fluorescence in situ
Hybridization (FISH)
Paraformaldehyde (PF)

ASJC Scopus subject areas

Anatomy
Histology

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10.1177/002215549804600811

Cite this

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title = "Optimized fixation and storage conditions for FISH analysis of single- cell suspensions",

abstract = "In our protocol to isolate and identify fetal cells in maternal peripheral blood, antibody (Ab)-stained cells are preserved with paraformaldehyde (PF) before batch flow cytometric sorting. However, PF fixation compromises the quality of subsequent interphase fluorescence in situ hybridization (FISH). We therefore examined the effect of PF concentrations and storage time in phosphate-buffered saline (PBS) on the quality of FISH signals. Cells were analyzed for changes in light scatter, morphology, and accessibility of target cell DNA. Fixation in 3\% PF for 1 hr was ideal for both flow cytometry and subsequent FISH detection. However, beyond 10 days of storage, FISH quality deteriorated.",

keywords = "Cell fixation methods, Flow cytometry, Fluorescence in situ, Hybridization (FISH), Paraformaldehyde (PF)",

author = "Sarah Murrell-Bussell and Dianne Nguyen and Schober, \{Wendy D.\} and Jeffrey Scott and Simpson, \{Joe Leigh\} and Sherman Elias and Bischoff, \{Farideh Z.\} and Lewis, \{Dorothy E.\}",

year = "1998",

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doi = "10.1177/002215549804600811",

language = "English (US)",

volume = "46",

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T1 - Optimized fixation and storage conditions for FISH analysis of single- cell suspensions

AU - Murrell-Bussell, Sarah

AU - Nguyen, Dianne

AU - Schober, Wendy D.

AU - Scott, Jeffrey

AU - Simpson, Joe Leigh

AU - Elias, Sherman

AU - Bischoff, Farideh Z.

AU - Lewis, Dorothy E.

PY - 1998/8

Y1 - 1998/8

N2 - In our protocol to isolate and identify fetal cells in maternal peripheral blood, antibody (Ab)-stained cells are preserved with paraformaldehyde (PF) before batch flow cytometric sorting. However, PF fixation compromises the quality of subsequent interphase fluorescence in situ hybridization (FISH). We therefore examined the effect of PF concentrations and storage time in phosphate-buffered saline (PBS) on the quality of FISH signals. Cells were analyzed for changes in light scatter, morphology, and accessibility of target cell DNA. Fixation in 3% PF for 1 hr was ideal for both flow cytometry and subsequent FISH detection. However, beyond 10 days of storage, FISH quality deteriorated.

AB - In our protocol to isolate and identify fetal cells in maternal peripheral blood, antibody (Ab)-stained cells are preserved with paraformaldehyde (PF) before batch flow cytometric sorting. However, PF fixation compromises the quality of subsequent interphase fluorescence in situ hybridization (FISH). We therefore examined the effect of PF concentrations and storage time in phosphate-buffered saline (PBS) on the quality of FISH signals. Cells were analyzed for changes in light scatter, morphology, and accessibility of target cell DNA. Fixation in 3% PF for 1 hr was ideal for both flow cytometry and subsequent FISH detection. However, beyond 10 days of storage, FISH quality deteriorated.

KW - Cell fixation methods

KW - Flow cytometry

KW - Fluorescence in situ

KW - Hybridization (FISH)

KW - Paraformaldehyde (PF)

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JF - Journal of Histochemistry and Cytochemistry

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ER -

Optimized fixation and storage conditions for FISH analysis of single- cell suspensions

Abstract

Keywords

ASJC Scopus subject areas

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