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TIRF time lapse of myosin-GFP in a cell with a symmetric monopolar spindle from metaphase to anaphase (cell was depleted of Klp61F [kinesin-5] and BubR1 by RNAi). At anaphase, myosin-GFP concentrates at the cell perimeter and becomes depleted from the central regions of the adhered cortex. Slowly, myosin breaks symmetry and ultimately relocalizes to the middle. This video corresponds to the image in Fig. 6 and video 10 from J Cell Bio 186:727-738, 2009. Total internal reflection microscopy was performed using a microscope (Perfect-focus TE2000; Nikon) with a 100×, 1.45 NA objective (Nikon) and illumination from either a 488-nm argon laser (100 mW) or a 491-nm solid-state laser (100 mW) and a 561-nm solid-state laser (50 mW). For dual-color TIRF microscopy, we used a triplepass dichroic filter (z491/561/633rpc) and changed the emission filter (ET525/50 or ET595/50; both from Chroma Technology Corp.) with a filter wheel placed before the camera. In some cases, an excitation notch filter was used in the filter cube (NF01-405/488/561/635; Semrock, Inc.). Images were typically captured every 2-3 s with a 50-200ms exposure with an EM charge-coupled device camera (iXon; Andor Technology). The microscope was controlled and images were acquired using open source MicroManager software (

Biological Sources
NCBI Organism Classification
Drosophila melanogaster
Cell Type
epithelial cell
Cell Line
Cellular Component
myosin regulatory light chain
Biological Context
Biological Process
mitotic anaphase
mitotic metaphase
kinesin 5 knockdown
BubR1 knockdown
Ronald D. Vale
James A. Spudich
Eric R. Griffis
J Cell Bio 186:727-738, 2009
Digital Object Identifier (DOI)
Archival Resource Key (ARK)
Grouping This image is part of a group.
Image Type
recorded image
Image Mode
Parameters Imaged
fluorescence emission
Source of Contrast
distribution of a specific protein
Visualization Methods
Processing History
unprocessed raw data
Data Qualifiers
raw, unprocessed data
suitable for spatial measurements
Sample Preparation
living tissue
Relation To Intact Cell
dispersed cells in vitro
Spatial Axis Image Size Pixel Size
X 268px 0.037µm
Y 242px 0.037µm
Time 3 seconds