Technical Reference #1841

1841.

Widespread Genomic Instability Mediated by a Pathway Involving Glycoprotein Ibα and Aurora B Kinase Youjun Li, Fengfeng L. Xu, Jie Lu, William S. Saunders, and Edward V. Prochownik, Children's Hospital of Pittsburgh of th University of Pittsburgh Medical Center, Journal of Biological Chemistry, 285(1841), (2010)
Link To Paper

Keywords:
DNA Damage, Mitosis, Myc, Oncogene, p53, Aneuploidy, Cleavage Furrow Regression, Double-stranded Breaks, Endoreduplication,Tetraploidy

Materials & Methods:
Microscopy—For fixed cell imaging, stained coverslips were mounted and examined under an Olympus BX60 epifluorescence microscope with 100 oil immersion objectives. A Hamamatsu Argus-20 CCD camera was used to capture the images. Confocal microscopy was performed using either an Olympus Fluoview 1000 microscope or a Nikon Eclipse E800 microscope equipped with a Bio-Rad Radiance 2000 system. For live cell microscopy, 2 105 cells were seeded on 35-mm glass-bottomed Petri dishes (MatTek Corporation, Ashland, MA)and subjected to live cell imaging with or without transient transfection with the indicated plasmids. The cells were video recorded while being maintained at 37 °C with a moisturized warm air microscope chamber (Life Imaging Services, Reinach, Switzerland). Differential interference contrast microscopy and epifluorescence microscopy were performed on Nikon Eclipse TE2000-U inverted microscope with CoolsnapHQdigital camera (Roper Scientific Photometrics, Tuczon, AZ). The images were taken and analyzed using MetaMorph software (Molecular Devices, Sunnyvale, CA). Cleavage furrow regression was determined visually from differential interference contrast microscopy images recorded at sequential time points.

Microscopic Technique
Confocal Microscopy

Cell Type(s)
Human Foreskin Fibroblasts (HFFs)