Coolsnap k4

Quantification of lagging chromosomes was performed using 50–500 images of contiguous fields acquired in an IN Cell Analyzer 2000 microscope (GE Healthcare) with a Nikon 40×/0.95 NA Plan Fluor objective (binning 2 × 2), using a large chip CCD Camera (CoolSNAP K4) with a pixel array of 1,024 × 1,024 (2.7027 pixel/µm resolution). All early to late anaphase figures were classified regarding the presence or absence of lagging chromosomes. Any DAPI-positive material between the two chromosome masses, but distinguishably separated from them, was counted as lagging chromosomes. DNA bridges were excluded from this analysis.

Images were acquired in an IN Cell Analyzer 2000 microscope (GE Healthcare) with a Nikon 20x/0.45 NA Plan Fluor objective (binning 1X1), using a large chip CCD Camera (CoolSNAP K4) with a pixel array of 2048x2048 (7.40 μm² pixel). Image field of view (FOV) x-y for this objective is 0.76x0.76 mm. With these settings and an approximate 70–80% cell confluence, 1500–2000 macrophages per well were imaged (per 4 FOV), a number we consider reasonable for the quantifications as it is 15 to 20-fold higher than that used with manual counting [15 (link)–17 (link)]. The excitation and emission filters used to detect DAPI, Alexa Fluor 568 and HCS CellMask^™ were DAPI, Texas Red, and Cy5, respectively.