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Ablation of cytoskeletal filaments and mitochondria in live cells using a femtosecond laser nanoscissor

Nan Shen, Dabajyoti Datta, Chris B. Schaffer, Philip LeDuc, Donald E.Ingber, Eric Mazur

Mechanics and Chemistry of Biosystems (2005)

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Analysis of cell regulation requires methods for perturbingmolecular processes within living cells with spatial discrimination on the nanometer-scale. We present a technique for ablating molecular structures in living cells using low-repetition rate, low-energy femtosecond laser pulses. By tightly focusing these pulses beneath the cell membrane, we ablate cellular material inside the cell through nonlinear processes. We selectively removed sub-micrometer regions of the cytoskeleton and individual mitochondria without altering neighboring structures or compromising cell viability. This nanoscissor technique enables non-invasive manipulation of the structural machinery of living cells with several-hundred-nanometer resolution. Using this approach, we unequivocally demonstrate that mitochondria are structurally independent functional units, and do not form a continuous network as suggested by some past studies.

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Femtosecond laser-drilled capillary integrated into a microfluidic device

Tyson N. Kim, Kyle Campbell, Alex Groisman, David Kleinfeld, and Chris B. Schaffer

Applied Physics Letters (2005)

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Recent growth in microfluidic technology is, to a large extent, driven by soft lithography, a high-throughput fabrication technique where polymer materials, such as polysdimethyld siloxane sPDMSd, are molded to form microscopic channel networks. Nevertheless, the channel architectures that can be obtained by molding are limited. We address this limitation by using femtosecond laser micromachining to add unmoldable features to the microfluidic devices. We apply laser ablation to drill microcapillaries, with diameters as small as 0.5 mm and aspect ratios as high as 800:1, in the walls of molded PDMS channels. Finally, we use a laser-drilled microcapillary to trap a polystyrene bead by suction and hold it against a shear flow

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Morphology of femtosecond laser-induced structural changes in bulk transparent materials

Chris B. Schaffer, Alan O. Jamison, and Eric Mazur

Applied Physics Letters (2004)

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Using optical and electron microscopy, we analyze the energy and focusing angle dependence of structural changes induced in bulk glass by tightly focused femtosecond laser pulses. We observe a transition from small density variations in the material to void formation with increasing laser energy. At energies close to the threshold for producing a structural change, the shape of the structurally changed region is determined by the focal volume of the objective used to focus the femtosecond pulse, while at higher energies, the structural change takes on a conical shape. From these morphological observations, we infer the role of various mechanisms for structural change

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All-Optical Histology Using Ultrashort Laser Pulses

Philbert S. Tsai, Beth Friedman, Agustin I. Ifarraguerri, Beverly D. Thompson, Varda Lev-Ram, Chris B. Schaffer, Qing Xiong, Roger Y. Tsien, Jeffrey A. Squier, and David Kleinfeld

Neuron (2003)

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As a means to automate the three-dimensional histological analysis of brain tissue, we demonstrate the use of femtosecond laser pulses to iteratively cut and image fixed as well as fresh tissue. Cuts are accomplished with 1 to 10 uJ pulses to ablate tissue with micron precision. We show that the permeability, immunoreactivity, and optical clarity of the tissue is retained after pulsed laser cutting. Further, samples from transgenic mice that express fluorescent proteins retained their fluorescence to within microns of the cut surface. Imaging of exogenous or endogenous fluorescent labels down to 100um or more below the cut surface is accomplished with 0.1 to 1 nJ pulses and conventional two-photon laser scanning microscopy. In one example, labeled projection neurons within the full extent of a neocortical column were visualized with micron resolution. In a second example, the microvasculature within a block of neocortex was measured and reconstructed with micron resolution.

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Bulk heating of transparent materials using a high-repetition-rate femtosecond laser

Chris B. Schaffer, Jose F. Garcia, and Eric Mazur

Applied Physics (2003)

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Femtosecond laser pulses can locally induce structural and chemical changes in the bulk of transparent materials, opening the door to the three-dimensional fabrication of optical devices. We review the laser and focusing parameters that have been applied to induce these changes and discuss the different physical mechanisms that play a role in forming them. We then describe a new technique for inducing refractive-index changes in bulk material using a high-repetition-rate femtosecond oscillator. The changes are caused by a localized melting of the material, which results from an accumulation of thermal energy due to nonlinear absorption of the high-repetition-rate train of laser pulses.

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Customization of Poly(dimethylsiloxane) Stamps by Micromachining Using a Femtosecond-Pulsed Laser

Daniel B. Wolfe, Jonathan B. Ashcom, Jeremy C. Hwang, Chris B. Schaffer, Eric Mazur, and George M. Whitesides

Advanced Materials (2003)

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This work describes the use of focused, high-intensity light from a Ti:sapphire laser that generates femtosecond pulses to create topographical structure in a flat surface of poly(dimethylsiloxane) (PDMS), and the use of the PDMS surfaces patterned in surface bas-reliefis the material most widely used for printing and stamping in soft lithography, and a material widely used in microfluidic systems. The bas-relief patterns required in these applications are usually fabricated by casting PDMS against a complementary bas-relief pattern in photoresist, fabricated in turn by photolithography. This process works well, but is not applicable to the preparation of PDMS stamps required for all types of problems; printing on spherical surfaces is an example.

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Microexplosions in tellurite glasses

S.K. Sundaram, Chris B. Schaffer, and Eric Mazur

Applied Physics (2003)

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Femtosecond laser pulses were used to produce localized damage in the bulk and near the surface of baseline, Al2O3-doped and La2O3-doped sodium tellurite glasses. Single or multiple laser pulses were non-linearly absorbed in the focal volume by the glass, leading to permanent changes in the material in the focal volume. These changes were caused by an explosive expansion of the ionized material in the focal volume into the surrounding material, i.e. a microexplosion. The writing of simple structures (periodic array of voxels, as well as lines) was demonstrated. The regions of microexplosion and writing were subsequently characterized using scanning electron microscopy (SEM), energy-dispersive spectrometry (EDS) and atomic force microscopy (AFM). Fingerprints of microexplosions (concentric lines within the region and a concentric ring outside the region), due to the shock wave generated during microexplosions, were evident. In the case of the baseline glass, no chemistry change was observed within the region of the microexplosion. However, Al2O3-doped and La2O3-doped glasses showed depletion of the dopant fromthe edge to the center of the region of the microexplosions, indicating a chemistry gradient within the regions. Interrogation of the bulk- and lasertreated regions using micro-Raman spectroscopy revealed no structural change due to the microexplosions and writing within these glasses

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Dynamics of femtosecond laser-induced breakdown in water from femtoseconds to microseconds

Chris B. Schaffer, Nozomi Nishimura, Eli N. Glezer, AlbertM.-T. Kim, and Eric Mazur

Optics Express (2002)

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Using time-resolved imaging and scattering techniques, we directly and indirectly monitor the breakdown dynamics induced in water by femtosecond laser pulses over eight orders of magnitude in time. We resolve, for the first time, the picosecond plasma dynamics and observe a 20 ps delay before the laser-produced plasma expands. We attribute this delay to the electron-ion energy transfer time.

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Interaction of femtosecond laser pulses with transparent materials

Chris B. Schaffer

Harvard University Ph.D. Thesis (2001)

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An intense femtosecond laser pulse can have an electric field strength which approaches or even exceeds the strength of the electric field that holds valence electrons in a transparent material to their ionic cores. In this regime, the interaction between the laser pulse and the material becomes highly nonlinear. Laser energy can be nonlinearly absorbed by the material, leading to permanent damage, and the material’s nonlinear response to the laser field can, in turn, induce radical changes in the laser pulse itself. The nature of these nonlinear interactions, the changes produced in the material and to the laser pulse, as well as several practical applications are explored in this thesis. We measure the laser intensity required to damage bulk transparent materials and uncover the dominant nonlinear ionization mechanism for different laser wavelengths and material band gaps. Using optical and electron microscopy, we examine the morphology of the material changes induced by tightly-focused femtosecond laser pulses in bulk transparent materials, and identify several mechanisms by which material changes are produced. We show that a high repetition rate train of femtosecond laser pulses can provide a point source of heat located inside the bulk of a transparent material, an effect which no other technique can achieve. The mechanism for white-light continuum generation is uncovered through measurement of the laser wavelength, the material band gap, and the external focusing angle dependence of the continuum spectrum. Using a time-resolved imaging technique, we follow the dynamics of the laser-produced plasma over eight orders of magnitude in time, revealing picosecond time scale dynamics that have not been previously observed. Finally, we discuss applications in direct writing of optical waveguides and in sub-cellular laser surgery.

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Laser-induced breakdown and damage in bulk transparent materials induced by tightly focused femtosecond laser pulses

Chris B. Schaffer, Andre Brodeur, and Eric Mazur

Measurement Science and Technology (2001)

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Laser-induced breakdown and damage to transparent materials has remained an active area of research for four decades. In this paper we review the basic mechanisms that lead to laser-induced breakdown and damage and present a summary of some open questions in the field. We present a method for measuring the threshold intensity required to produce breakdown and damage in the bulk, as opposed to on the surface, of the material. Using this technique, we measure the material band-gap and laser-wavelength dependence of the threshold intensity for bulk damage using femtosecond laser pulses. Based on these thresholds, we determine the relative role of different nonlinear ionization mechanisms for different laser and material parameters.

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