Centrosome Size Assay

Quantitative and unbiased characterization of centrosome growth in 3D time-lapse images requires automated methods. We developed tools for tracking and measuring the size of fluorescently labeled centrosomes as well as detecting the embryo contour, naming cells and detecting the nuclear envelope breakdown in each cell division event. This allows us to convert raw images into statistics of centrosome growth for wild-type and mutant embryos with minimal manual effort. Validated with beads injected into embryos we found our measurements of object radius to be within 50 nm of the true radius.

Quantifying centrosome growth over multiple rounds of cell division in wild-type embryos expressing ϒ-Tubulin::GFP imaged with a spinning disc confocal microscope at 96x magnification and time-lapse interval of 40 seconds.

Measuring the effect of cell size on centrosome size using 2-cell stage wild-type and ani-2(RNAi) embryos, both expressing ϒ-Tubulin::GFP imaged with a spinning disc confocal microscope at 96x magnification and time-lapse interval of 20 seconds.