Single-cell nanosurgery

Nanosurgery, as applied in biology, involves the use of laser beams focused by an objective microscope lens to exert a controlled force onto, and therefore manipulate, organelles and other subcellular structures. The technique, which is so precise that it allows for the destruction of a single cell without damaging adjacent ones, may eventually revolutionize molecular cell biology.

Conventional nanosurgery uses optical tweezers, consisting of beams of laser light, to manipulate dielectric particles (that is, insulators, or particles which are highly resistant to electric currents). The narrowest point of the laser beam contains a strong electric field gradient, which attracts dielectric particles such that they move along the gradient towards the centre of the beam, where the electric field is strongest. The attraction of the dielectric particles to the centre of the laser beam enables the particles to be moved from one location to another, without ever touching them.

The process by which particles are attracted to the laser beam is called ‘trapping’, and was first discovered by Arthur Ashikin of Bell Laboratories in 1970. It was originally developed as a technique for manipulating atoms – atomic physicists can use beams of focused light to cool atoms to extremely low temperatures – but optical tweezers can also applied to biological structures such as cell nuclei and chromosomes. Again, it was Ashkin and his colleagues who first demonstrated the use of optical tweezers on biological materials – used to trap individual tobacco mosaic virus particles and Escherichia coli bacteria. Since then, the technique has been used to investigate the mechanical properties of microtubules and to measure the forces generated by individual molecules of motor proteins.

The use of optical tweezers in combination with a laser scalpel allows for even greater precision. For example, researchers at Eric Mazur‘s photonics laboratory at Harvard University used used femtosecond pulses of light of a near-infrared wavelength to ablate AFD sensory neurons in the nematode worm Caenhorhabditis elegans (where a femtosecond is one quadrillionth of a second). Similarly, Adela Ben-Yakar and her colleagues at the University of Austin in Texas, used femtosecond pulses to sever individual motor axons in the nematode worm, in order to study nerve regeneration. Once the apparatus was set up, each procedure using the optical ‘nano-scissors’ took about ten minutes to perform.

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D-type motor neurons in the nematode worm before and after laser ablation with femtosecond pulses of near-infrared light (from Yanik, M. F. et al, 2006)

Nanosurgery does, however, have its drawbacks. Many researchers engineer their organisms to fluoresce, so that the cells being investigated can be visualized easily. When a structure is trapped during nanosurgery, it is attracted to the centre of the laser beam, and because light intensity is greatest at the centre of the beam, photobleaching – the loss of fluorescence – occurs, which damages the trapped structure.

A recent paper in the journal Nano Letters describes a novel nanosurgical technique which does not cause photobleaching of trapped structures. The method involves the use of a polarization-shaped optical vortex trap. In other words, the light in the laser beam twists like a corkscrew around the axis of propagation. (This is known as a Laguerre-Gaussian, or LG, beam.) At the wavefront of the beam, light is therefore distributed in the shape of a helix. The wavefront has a ‘dark core’, at which light intensity is zero. During trapping, this dark core damages biological components far less than conventional optical tweezing techniques.

Daniel Chiu and his colleagues, at the University of Washington, who developed the polarization-shaped optical vortex trap, used it to trap individual mitochondria within living cells. They also measured the time course over which the organelles underwent photobleaching, and found that, whereas mitochondria trapped in optical tweezers were bleached within 2 seconds, those trapped in an optical vortex remained stable over the same period of time.

Nanosurgery has many applications in the biological sciences. It could, for example, be very useful to researchers investigating the development of the nervous system. During certain stages of neural development, trail-blazing ‘pioneer’ neurons migrate and provide chemical signals which subsequently migrating cells use to extend their processes in the right way. Nanosurgery could be used to eliminate pioneer neurons in order to determine exactly how they influence cells which.

The method may also have clinical applications. According to Professor Chiu:

The ability to manipulate optically submicrometer subcellular structures while minimizing photodamage has several practical applications. In particular, we are developing a droplet nanolab platform for single-organelle assays and chemical analysis, because many diseases are caused by organelle malfunction where a given cell can contain a heterogeneous population of healthy and defective organelles.

References:

Jeffries, G. D. M. (2006) Using Polarization-Shaped Optical Vortex Traps for Single-Cell Nanosurgery Nano Lett. DOI: 10.1021/nl0626784

Ashkin, A. (1997). Optical trapping and manipulation of neutral particles using lasers. Proceedings of the National Academy of Sciences 94: 4853-4860.

2 thoughts on “Single-cell nanosurgery

  1. I had the good fortune of visiting Dr. Ben-Yakar’s lab. Her setup is quite impressive (not to mention the results). Hopefully our lab will collaborate with hers at some point in the near future. We do all our transections with sharpened glass pipettes, certainly not as elegant as the femtosecond laser.

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