Eirini Papagiakoumou

Light-induced cell separation in a tailored optical landscape

We demonstrate passive optical sorting of cell populations in the absence of any externally driven fluid flow. Specifically, we report the movement of erythrocytes and lymphocytes in an optical landscape, consisting of a circularly symmetric light pattern created by a Bessel light beam. These distinct cell populations move, spontaneously and differentially, across the underlying periodic optical landscape. Thus, we were able to separate lymphocytes from a mixed population of cells containing erythrocytes and then collect the lymphocytes in a microcapillary reservoir. We also demonstrate an enhanced form of this separation that exploits the polarizability of silica microspheres by attaching spheres coated with antibodies to cell surface markers to a subpopulation of lymphocytes. These techniques may be applied using standard laboratory apparatus.

Passive optical separation within a 'nondiffracting' light beam

A passive, optical cell sorter is created using the light pattern of a nondiffracting beam-the Bessel beam. As a precursor to cell sorting studies, microspheres are used to test the resolution of the sorter on the basis of particle size and refractive index. Variations in size and, more noticeably, refractive index, lead to a marked difference in the migration time of spheres in the Bessel beam. Intrinsic differences (size, refractive index) between native (unlabeled) cell populations are utilized for cell sorting. The large difference in size between erythrocytes and lymphocytes results in their successful separation in this beam pattern. The intrinsic differences in size and refractive index of other cells in the study (HL60 human promyelocytic leukaemic cells, murine bone marrow, and murine stem/progenitor cells) are not large enough to induce passive optical separation. Silica microsphere tags are attached to cells of interest to modify their size and refractive index, resulting in the separation of labeled cells. Cells collected after separation are viable, as evidenced by trypan blue dye exclusion, their ability to clone in vitro, continued growth in culture, and lack of expression of Caspase 3, a marker of apoptosis.

Q-switched Versus Free-running Er:YAG Laser Efficacy on the Root Canal Walls of Human Teeth: A SEM Study

Twenty-one teeth with one root canal were prepared by the step-back technique, divided into three groups, and split longitudinally. Group A served as a control. In group B, 20 to 150 pulses of 100 micros, 30 to 70 mJ per pulse at 1 to 4 Hz from a free-running Er:YAG laser were applied to the root-canal dentin. In group C, the Q-switched Er:YAG laser, with the same energy parameters and a 190-ns pulse duration was used. Scanning electron microscopy examination revealed that control specimens had debris and smear layer obscuring the dentinal tubules at all levels in the canals without crack formation. Both groups of laser-treated dentin were clean with opened dentinal tubules except around the lased area in which there was an intact smear layer. Cracks were observed in both laser groups with higher frequency in group C. In group B, craters with different depth levels at the root canal walls were produced and the energy apparently was distributed equally, because craters were well-shaped. In contrast, the ablation efficiency in group C was questionable with the parameters used in this study. Consequently, suitable parameters of the free-running Er:YAG laser must be found before its careful use as an adjunct in endodontic therapy.

Evaluation of trapping efficiency of optical tweezers by dielectrophoresis

A relatively new method for measuring optically induced forces on microparticles and cells, different from the conventional Brownian motion and viscous drag force calibration methods widely used, is introduced. It makes use of the phenomenon of dielectrophoresis for the calibration of optical tweezers through the dielectrophoretic force calculations. A pair of microelectrodes is fabricated by photolithography on a microscope slide and it is connected to a high-frequency generator. The calibration of the optical tweezers setup is performed by the manipulation of polystyrene beads and yeast cells. Calibration diagrams of the transverse forces versus power are deduced for different cell radii and numerical apertures of the objective lenses. The optical system and the related technique provide a fast and easy method for optical tweezers calibration.

Comparative evaluation of HP oxide glass fibers for Q-switched and free-running Er:YAG laser beam propagation

The radiation transmission of a Q-switched and a free-running Er:YAG laser, emitting at 2.94 μm, through high power (HP) oxide glass fibers of 450 μm and 250 μm core diameter was studied. Attenuation measurements were obtained as a function of the laser energy input and as a function of the curvature. The output beam quality was also studied using a beam profiler. Experiments with the same Er:YAG laser but with a 1000 μm core diameter cyclic olefin polymer coated silver hollow glass (COP/Ag) waveguide as the delivery system were performed for comparison. The results are promising as far as the delivery of Q-switched Er:YAG laser radiation is concerned. The fibers exhibited an attenuation below 0.7 dB/m, good output beam profiles, while no damage was observed after extended use.

Q-switched Er:YAG radiation transmission through an oxide glass fiber for medical applications

In the last few years, there has been an increasing interest for the 3 μm laser radiation in various medical applications, as this wavelength is strongly absorbed by the water and the other components of soft and hard tissue. An intensive development effort is going on throughout the world, in order to develop reliable lasers emitting in the 3 μm wavelength range. Our laser development effort with the Q-switched Er:YAG laser is briefly described in this article. Additionally for medical applications there is a great demand for good flexible delivery systems, in the mid-IR wavelength region. In this work the radiation transmission of a Q-switched Er:YAG laser, emitting at 2.94 μm, through high power (HP) oxide glass fibers of 450 μm core diameter was studied. Attenuation measurements were obtained as a function of the laser energy input and as a function of curvature, at 90 °, 180° and 360° bending angle. The output beam quality was studied using a beam profiler. Experiments with the same delivery system transmitting free-running Er:YAG laser radiation, were performed for comparison. The results are promising for the delivery of Q-switched Er:YAG laser radiation, as the fibers exhibited attenuation of 0.7 dB/m, and no damage of them was observed.

Cell sorting in a static optical potential landscape

Microscopic particles with varying optical properties may be induced to move in different ways when placed on a sculpted optical potential due to differences in shape, size or polarisability. The separation of red blood cells (erythrocytes) and white blood cells (lymphocytes) is achieved in a non-invasive manner and in the absence of any microfluidic systems using a non-diffracting circularly symmetric Bessel beam. The Bessel beam, which consists of a series of concentric rings, each of equal power and of 3.2μm thickness with a spacing of 2μm around a central maximum of 5μm diameter (and is akin to a rod of light as its propagation distance is 3mm), is directed upward into a sample chamber containing blood. Fluctuations in Brownian motion cause cells to escape from individual rings of the Bessel beam and travel towards the beam centre, where the intensity of the rings increases. However, these cells must be able to overcome the potential barrier of each ring which gets larger toward the central maximum. Lymphocytes - spherical in shape and 7μm in diameter (therefore overlapping two rings) - are transported, due to the gradient force of the optical field, to the beam centre where they are guided upwards and form a vertical stack, whereas erythrocytes re-align on their sides in the outer rings and are then guided upwards, because once aligned they cannot escape the potential barrier and lock-in to that ring. The optical power required for optimal sorting in this static sorter which requires no fluid flow is investigated.

Q-switched Er:YAG radiation transmission through medical sapphire fibers

The effect of the 2.94 μm Er:YAG laser radiation propagation through sapphire fibers with diameters varying from 250 μm to 550 μm, on the quality of the laser beam is investigated. A comparison was made between the fibers performance in free-running and Q-switched Er:YAG laser radiation.

Determination of the maximum capabilities of high-power oxide glass fibers in the mid-infrared region for medical applications

An intensive development effort is going on throughout the world, in order to develop reliable lasers emitting in the 3 μm wavelength range, as this wavelength is strongly absorbed by the water and the other components of soft and hard tissue and thus its use is important in various medical applications. In parallel, good flexible delivery systems, in the mid-IR wavelength region, are needed in order to deliver the laser beam to the tissue. In this work High Power (HP) Oxide Glass fibers are tested for determining their maximum capabilities in delivering free-running and Q-switched Er:YAG laser radiation at 2.94 μm. Oxide glass is a new material in solid core fiber fabrication for medical applications, and its performance at the wavelength of 2.94 μm, for various laser characteristics is of great importance. Also a comparison is made between results obtained with the two different Er:YAG lasers, afree-running and a Q-switched one, and the results obtained at 2.78 μm, with a chemical HF laser.

Q-switched Er:YAG radiation transmission through medical COP-coated silver hollow glass waveguide

Transmission measurements of Q-switched Er:YAG laser radiation, through cyclic olefin polymer-coated silver hollow glass waveguides, were performed under straight and bent conditions and the beam quality at the output of the waveguide was studied.