PMC:6080766 / 8076-9405 JSONTXT

{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/6080766","sourcedb":"PMC","sourceid":"6080766","source_url":"https://www.ncbi.nlm.nih.gov/pmc/6080766","text":"Sister Chromatid Exchange Assay\nDermal fibroblasts were treated with 10 μM BrdU for 48 hr followed by 0.5 μg/mL colcemid for 2 hr. Metaphases and nuclei were isolated in hypotonic buffer (0.25% KCl and 1% Na3C6H5O7), fixed with methanol and acetic acid (3:1 vol/vol), and dropped onto slides. Dried slides were rehydrated in PBS and then incubated with 2 μg/mL Hoescht 33342 stain in 2× saline sodium citrate (SSC) buffer (300 mM NaCl and 30 mM sodium citrate) for 15 min. Slides were then covered in 2× SSC buffer, irradiated in ultraviolet A at 5,400 J/m2, dehydrated in an ethanol series, and mounted in VECTASHIELD with DAPI.\n\nSCE Methodology for Analysis of P7 and P8\nDermal fibroblasts were treated with 26.05 μM BrdU for 72 hr followed by 0.5 μg/mL colcemid for 5 hr and then harvested. Cells were resuspended in a pre-warmed hypotonic solution (0.051 M KCl) for 20 min at 37°C, fixed with methanol and acetic acid (3:1 vol/vol), and dropped onto slides. Samples were aged for at least 3 days and then stained with 1.56 μg/mL Hoechst 33258 (Sigma, B2883). Slides were irradiated in 2× SSC with ultraviolet C (356 nm) at 0.260 J/cm2, washed with 2× SSC, and stained with 10% Giemsa in phosphate buffer.\nSCE analysis of peripheral-blood samples was performed according to standard methodology in a diagnostic lab setting.27\n","divisions":[{"label":"Title","span":{"begin":0,"end":31}},{"label":"Section","span":{"begin":631,"end":1328}},{"label":"Title","span":{"begin":631,"end":672}}],"tracks":[]}