Slides of metaphase spreads were stained with quinacrine mustard (Sigma-Aldrich) and Hoechst 33258 (Sigma-Aldrich) for observation of the chromosome size and banding pattern. Metaphase images were captured digitally with a CoolCubeI CCD camera mounted on a fluorescence microscope were processed using the ISIS software provided with the microscope.
Quinacrine mustard
Manufactured by Merck Group
Quinacrine mustard is a chemical compound used in laboratory applications. It is a synthetic dye that exhibits fluorescent properties. The primary function of quinacrine mustard is as a staining agent for the visualization and analysis of cellular structures in microscopy techniques.
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Lab products found in correlation
3 protocols using quinacrine mustard
1
Metaphase Chromosome Analysis
2
Immunofluorescent Labeling of Histone Modifications
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After the transfection procedure cells were permeabilized with 0.5% Triton X100 for 5 min at room temperature. Cells were then briefly washed several times with PBS, and unspecific binding was reduced by incubation in PBSB (PBS supplemented with 1% bovine serum) for 20 min at 37 °C in a moist chamber. Subsequently, cells were immersed in primary antibodies diluted in PBSB for 1 h at 37 °C. Thereafter cells were washed 3× 30 min in PBST (PBS supplemented with 0.05% Tween20) and incubated in secondary antibodies diluted in PBSB for 30 min at 37 °C. Cells were washed 3× 30 min in PBST, counterstained with DAPI (Sigma) or quinacrine mustard (Sigma), and coverslips were mounted in Citifluor antifading mounting solution (Agar Scientific).
Primary antibodies used were: monoclonal anti- HIRA68 (link) (1:10; mixture of monoclonal antibodies WC19, WC117, WC119); rabbit polyclonal anti-H3K27me3 (1:1000; Upstate 07–449); monoclonal anti- SC-35 (1:2000; Sigma S4045); monoclonal anti- H3K4me2 and -me3 -specific (1:500; Abcam ab6000). Secondary antibodies were goat anti-mouse Alexa488 and goat anti-rabbit Alexa488 (1:1000; Life Technologies).
Primary antibodies used were: monoclonal anti- HIRA68 (link) (1:10; mixture of monoclonal antibodies WC19, WC117, WC119); rabbit polyclonal anti-H3K27me3 (1:1000; Upstate 07–449); monoclonal anti- SC-35 (1:2000; Sigma S4045); monoclonal anti- H3K4me2 and -me3 -specific (1:500; Abcam ab6000). Secondary antibodies were goat anti-mouse Alexa488 and goat anti-rabbit Alexa488 (1:1000; Life Technologies).
3
Chromosome Staining for Mitotic Analysis
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Colcemid solution (KaryoMAX™, Thermo Fisher Scientific) was added overnight to cultured cells in logarithmic phase. Chromosomes were stained with the fluorescent dye quinacrine mustard (Sigma‐Aldrich). Q‐Band stained chromosomes were analysed.
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