Figure 1.

The optical setup used in the laser manipulation of early to mid cleavage stage (2 to 4/8 cell) zebrafish embryos. Sub-10 fs laser pulses were generated from a Kerr lens modelocked titanium sapphire laser oscillator. The centre wavelength of the laser pulses was at 800 nm, with a pulse repetition rate of 80 MHz. Fs laser pulses were focused by a 1.0 NA water immersion microscope objective to a focal spot of ~800 nm. A galvanometer was used to select the beam dwell time irradiating the embryos. The embryos were placed on a motorized x-y-z stage, with an in plane translation speed of 1 mm/sec and a z-focus step resolution of 50 nm. White light illuminated the samples in the inverted position. For fluorescence assessment, fluorescence excitation was collinearly coupled through the objective onto the sample. A charge-coupled device (CCD) was used to observe the laser-manipulation of the embryos. The CCD was interfaced with a computer, allowing for video capture and image analysis.