Free Ebook Laser tweezers for moving live dissociated neurons

A laser tweezers system for transporting dissociated neurons into small “cages” on a culture dish was constructed, and it was studied extensively.

The system consists of an inverted microscope, a 1064 nm or 980 nm laser module, a beam expander, a motorized mechanical stage, a CCD camera, and steering mirrors. A laser beam is generated by the IR laser module, and the beam is expanded by the beam expander to match the size of the back aperture of the objective. The beam is then steered into the objective where it is focused to a point. The system uses this single, tightly focused laser beam to trap a neuron. Once a neuron is trapped and lifted, the mechanical stage is moved to locate the neuron above its destination. The system will know the location of the neurocages and will automatically move neurons to their destination.

Newly dissociated neurons will attach to most substrate surfaces eagerly, and the lifting of a neuron is impossible when it is attached to the surface. Many possible surfaces were investigated, and it was discovered that the surface can best be made “non-sticky” for more than an hour, by coating the surface with Poly-2-hydroxyethyl methacrylate (PolyHEMA). The neural survival at different laser intensities, exposure times, and wavelengths were studied. The results show that neural survival depends strongly on laser wavelengths, and a 980 nm laser is less damaging than a 1064 nm laser. For 980 nm, perfect survival after irradiation is independent of laser power up to our maximum of 130 mW for exposure time up to 4 minutes. At 17 mW, almost all neurons can be lifted off a PolyHEMA substrate. The maximum speed for moving a neuron through the medium at different laser intensities was studied, and was 250 um/s at 100 mW for 980 nm. The studies have shown that a laser tweezers system is suitable for transporting live dissociated neurons over millimeter distance in less than a minute. The neural survival in neurocages on glass substrate was then studied. The survival and growth over time for neurons loaded into cages was found to be no different than for that of a control culture.

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