Abstract
We demonstrate a system for the combined optical injection and trapping of developing embryos. A Ti:sapphire femtosecond laser in tandem with a spatial light modulator, is used to perform fast and accurate beam-steering and multiplexing. We show successful intracellular delivery of a range of impermeable molecules into individual blastomeres of the annelid Pomatoceros lamarckii embryo by optoinjection, even when the embryo is still enclosed in a chorion. We also demonstrate the ability of the femtosecond laser optoinjection to deliver materials into inner layers of cells in a well-developed embryo. By switching to the continuous wave mode of the Ti:sapphire laser, the same system can be employed to optically trap and orient the 60 μm sized P. lamarckii embryo whilst maintaining its viability. Hence, a complete all-optical manipulation platform is demonstrated paving the way towards single-cell genetic modification and cell lineage mapping in emerging developmental biology model species.
Highlights
Optical manipulation allows contact-free handling [1] and modification of microscopic biological samples
16 May 2011 1 June 2011 / Vol 2, No 6 / BIOMEDICAL OPTICS EXPRESS 1565 same time a pulsed focused laser beam of sufficient intensity can porate the membrane of a single cell leading to optical injection of molecules and genetic material [3,4]
A single femtosecond laser system can be toggled between continuous wave (CW) and fs operation for optical trapping of cells and intracellular delivery of macromolecules [6], as we show in this paper
Summary
Optical manipulation allows contact-free handling [1] and modification of microscopic biological samples. Optical trapping of a variety of swimming motile specimens was reported using a dual focus mirror trap [1] These results show that a non-contact automated optical method to move, orient and hold developing embryos would bring a clear advantage over the commonly used intrusive glass capillaries, which cause unnecessary stress in the sample and require manual dexterity,. By toggling between CW and pulsed modelocked operation, we demonstrate independent optical trapping of the 60 μm sized embryos of Pomatoceros lamarckii and optical injection of macromolecules into its individual blastomeres. By changing the light wavefront modulation encoded on the SLM, three dimensional beam steering and multiplexing can be achieved Using this system, individual embryos can be positioned and oriented in 3-dimensions using a low numerical aperture (NA.) objective, allowing optical orientation and manipulation within a large-field of view. The versatility and ease-of-use offered by this combined system opens new avenues in flexible and dynamic manipulation of developing embryos
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