An optical scalpel and optical tweezer have been combined to perform intracellular microsurgery and micromanipulation in vivo. When only laser microsurgery was performed on metaphase chromosomes, the dissected sister chromatid fragments drifted off to either the side of the spindle or completely off the spindle. At anaphase the fragments separated and the two arms generally moved to their respective daughter cells. When the chromosome arm was cut during anaphase A and B, the distal chromosome fragment separated from the rest of the chromosome and moved toward the pole, following the proximal chromosome fragment. Distal chromosome fragments laser-dissected during metaphase were held together throughout anaphase using the optical trap. Optical trapping of dissected chromosome fragments during anaphase A and B inhibited movement of the chromosome fragment to its pole. As a result, the trapped chromosome fragments were (1) incorporated into the opposite daughter cell, (2) lost in the cleavage furrow during cytokinesis, or (3) eventually incorporated into the correct daughter cell. These results indicate that optical traps are effective in holding laser-dissected chromosome fragments throughout mitosis. This new tool should be useful for studies on chromosome movement and cell genetics.

The chick chorioallantoic membrane (CAM) model was used to study vascular effects of photodynamic therapy (PDT) and hyperthermia (HPT) and the synergism of these modalities. The CAM is a convenient medium for monitoring the modifications of the vasculature. It is possible to view the CAM and to examine structural changes of individual blood vessels in real tune. Moreover, the CAM is a closed system which lends itself to mathematical modeling of the temporal and spatial temperature profile and in which HPT can be performed quantitatively and to a selected depth, using different lasers. A porphyrin‐type photosensitizer solution was applied to areas of the CAM, defined by teflon O‐rings placed on the surface. Uptake dynamics of the sensitizer into the CAM was determined by analyzing its fluorescence in vivo. The CAM area was irradiated with a dual‐wavelength laser system composed of a dye laser at 644 nm (to induce PDT) and a CO 2 laser at 10.6 μm (to bring about HPT). Damage to the CAM vasculature, due to combined PDT + HPT, was compared to the outcome of the separate modalities, and a synergistic effect of about 40% was observed. © 1992 Wiley‐Liss, Inc.

A single-beam gradient force optical trap was combined with a pulsed UV laser microbeam in order to perform laser induced cell fusion. This combination offers the possibility to selectively fuse two single cells without critical chemical or electrical treatment. The optical trap was created by directing a Nd:YAG laser, at a wavelength of 1.06 microns, into a microscope and focusing the laser beam with a high numerical aperture objective. The UV laser microbeam, produced by a nitrogen-pumped dye laser (366 nm), was collinear with the trapping beam. Once inside the trap, two cells could be fused with several pulses of the UV laser microbeam, attenuated to an energy of approximately 1 microJ/pulse in the object plane. This method of laser induced cell fusion should provide increased selectivity and efficiency in generating viable hybrid cells.

A single-beam gradient force optical trap was combined with a pulsed UV laser microbeam In order to perform laser Induced cell fusion. This combination offers the possibility to selectively fuse two single cells without critical chemical or electrical treatment. The optical trap was created by directing a Nd:YAG laser, at a wavelength of 1.06 jim, into a microscope and focusing the laser beam with a high numerical aperture objective. The UV laser microbeam, produced by a nitrogenpumped dye laser (366 rim), was collinear with the trapping beam. The maximum transverse force exerted on a moving cell (NS- 1) by the optical trap was determined by measuring the velocity at which the cell fell out of the trap. For a laser power of 55 mW and a cell diameter from 9 to 20 rim, the drag force was calculated using Stokes' law to be in the range from 1.42 0.22 x 106 to 1.13 0.25 x 10-6 dynes. For the fusions, the transverse force needed to capture two NS- 1 cells and to bring them into close contact, was > 200 mW. Once inside the trap, two cells could be fused with several pulses of the UV laser microbeam, attenuated to an energy of 1 &J/pulse In the object plane. This method of laser Induced cell fusion should provide Increased selectivity and efficiency in generating viable hybrid cells.