Laser-induced cavitation fascinates because it involves a sequence of nonlinear interactions (plasma formation, shock wave emission, and bubble dynamics) but the experimental investigation is challenging due to the large range of spatial and temporal scales. We imaged laser and bubble interactions with a solid target during laser ablation in liquids in side view using a water immersion microscope objective with high numerical aperture. For stroboscopic and high-speed imaging, femtosecond laser pulses were coupled into a multimode fiber. With optimized fiber length, spatial mode scrambling provides speckle-free illumination that enabled us to freeze bubble and shock wave dynamics at diffraction-limited resolution.
Laser-induced cavitation accompanies laser surgery in cells and transparent tissues and its engineering can improve surgical results. To elucidate the underlying mechanisms, plasma-mediated shock wave formation and bubble dynamics are investigated by stroboscopic and high-speed photography with ultrahigh spatiotemporal resolution. We developed a novel light source for speckle-free illumination at exposure times < 100 ps that is based on amplified spontaneous emission (ASE) and lasing in a femtosecond-laser-pumped Rhodamine dye cell. The contributions from ASE and lasing and their influence on pulse duration, divergence and coherence are investigated, and the emission characteristics are optimized for high repetition rates.
Pulsed laser ablation in liquids (LAL) is a growing field for the generation of metallic nanoparticles, but a comprehensive picture of the fundamental mechanisms is still missing. Material ejection, shock wave, and cavitation bubble dynamics are investigated by high-speed photography with speckle-free illumination at exposure times below 100 ps using a novel light source that is based on amplified spontaneous emission within a femtosecond laser pumped Rhodamine dye cell. Framing rates at up to 1 MHz and accurate triggering even make it possible to capture the bubble collapse at LAL.
Pulsed laser ablation in liquids (LAL) is a growing field for the generation of metallic nanoparticles, but a comprehensive picture of the fundamental mechanisms is still missing. The process is always associated with rapid material ejection, shock waves and cavitation. Time-resolved photography with speckle-free illumination at exposure times below 200 ps can “freeze” these events and provide diffraction-limited resolution. We present a novel light source fulfilling these demands based on a SBS compressed Q-switched laser pulse pumping a dye cell and illustrate it’s utility through photography of LAL events on a gold target.
Photodamage in nonlinear microscopy of transparent tissues starts at irradiances 1.5 times above the autofluorescence imaging level. Although the free-electron density is low, their energy suffices to break bonds in water, DNA and the backbone and side chains of proteins. We explored photodamage kinetics using physical indicators (hyperfluorescence, plasma luminescence, bubble formation). By plotting threshold values in (irradiance/radiant exposure) space, we identified a “safe” region for microscopy. Thermomechanical effects become relevant in melanin-containing tissue. Two-photon excitation of retinal fluorophores allows monitoring metabolic transformations. We analyze the thermomechanical damage pathways in retinal imaging, and discuss strategies for mitigating such damage.
Laser-induced plasma generation by single and multiple femtosecond laser pulses is used surgically and constitutes a source of photodamage in nonlinear microscopy. The irradiance threshold at which transient vapor bubbles in water are produced is 20x higher than the irradiance used for nonlinear microscopy. However, photodamage in multiphoton microscopy already starts, when the irradiance is 1.5x above autofluorescence imaging. Thus, there is a huge realm of low-density plasma effects between the multi-pulse damage threshold and the single-pulse surgical regime, and the talk will provide a systematic overview over laser applications and the irradiance and radiant exposure dependence of these effects.
Femtosecond laser-induced plasma generation is used surgically and may also cause photodamage in nonlinear microscopy. Photodamage in multiphoton microscopy already starts at irradiances 1.5 times above the value used for autofluorescence imaging but the cavitation bubble threshold is 20 x higher. We explore the realm of low-density plasma effects between multi-pulse nonlinear imaging and single-pulse surgical regime. We characterize the transition from unchanged tissue (emitting autofluorescence) to slightly changed tissue (hyperfluorescence), drastically changed tissue (plasma luminescence) and disintegrated biomolecules (gas bubble formation). By plotting the threshold values in (irradiance, radiant exposure) space, we identified a “safe” region for nonlinear microscopy.
Correction of hyperopia requires an increase of the refractive power by steepening of the corneal surface. Present refractive surgical techniques based on corneal ablation (LASIK) or intrastromal lenticule extraction (SMILE) are problematic due to epithelial regrowth. Recently, it was shown that correction of low hyperopia can be achieved by implanting intracorneal inlays or allogeneic lenticules. We demonstrate a steepening of the anterior corneal surface after injection of a transparent, liquid filler material into a laser-dissected intrastromal pocket. We performed the study on ex-vivo porcine eyes. The increase of the refractive power was evaluated by optical coherence tomography (OCT). For a circular pocket, injection of 1 μl filler material increased the refractive power by +4.5 diopters. An astigmatism correction is possible when ellipsoidal intrastromal pockets are created. Injection of 2 μl filler material into an ellipsoidal pocket increased the refractive power by +10.9 dpt on the short and +5.1 dpt on the long axis. OCT will enable to monitor the refractive change during filler injection and is thus a promising technique for real-time dosimetry.
Understanding free-electron mediated effects of tightly focused femtosecond pulse series is essential for minimizing photodamage in nonlinear microscopy and opens new avenues for nanosurgery and intentional modifications of biomolecules. We tracked different stages of the photomodification kinetics (hyperfluorescence, plasma luminescence, bubble formation) by time-lapse 2-photon microscopy, fluorescence lifetime measurements, and bubble interferometry with nanometer resolution. Monitoring of bubble growth during pulse series enabled us to quantify chemical reaction rates leading to gas formation via molecular disintegration. Novel ways of data evaluation were used to create a comprehensive picture of the photomodification kinetics in the (irradiance/irradiation dose) parameter space.
For ametropic eyes, LASIK is a common surgical procedure to correct the refractive error. However, the correction of hyperopia is more difficult than that of myopia because the increase of the central corneal curvature by excimer ablation is only possible by intrastromal tissue removal within a ring-like zone in the corneal periphery. For high hyperopia, the ring-shaped indentation leads to problems with the stability and reproducibility of the correction due to epithelial regrowth.
Recently, it was shown that the correction of hyperopia can be achieved by implanting intracorneal inlays into a laser-dissected intrastromal pocket. In this paper we demonstrate the feasibility of a new approach in which a transparent, and biocompatible liquid filler material is injected into a laser-dissected corneal pocket, and the refractive change is monitored via OCT. This technique allows for a precise and adjustable change of the corneal curvature.
Precise cutting of the intrastromal pocket was achieved by focusing UV laser picosecond pulses from a microchip laser system into the cornea. After laser dissection, the transparent filler material was injected into the pocket. The increase of the refractive power by filler injection was evaluated by taking OCT images from the cornea. With this novel technique, it is possible to precisely correct hyperopia of up to 10 diopters. An astigmatism correction is also possible by using ellipsoidal intrastromal pockets.
Studying the wavelength dependence of femtosecond optical breakdown in water helps resolving an ongoing controversy on the relative importance of multiphoton, tunneling and avalanche ionization. Measurements of the bubble formation threshold at 50 wavelengths from UV to near-IR revealed a continuous decrease of the irradiance threshold with increasing wavelength. This is indicative for a dominant role of avalanche ionization, which gains strength with wavelength whereas the multiphoton ionization rate decreases.
Fitting data by a model considering breakdown initiation via a solvated electron state yielded an effective Drude electron collision time of 1 fs. Modeling predicts that the threshold continues to decrease up to 1.3 μm but levels out for longer wavelengths. It remains low in the mid IR because wavelength-independent tunneling ionization ensures a constant level of seed electrons for the ionization avalanche even though the influence of multiphoton ionization ceases.
The low breakdown threshold opens promising perspectives for ultrashort-pulsed laser surgery at wavelengths around 1.3 μm and 1.7 μm, which are attractive due to a favorable combination of low scattering and moderate water absorption. The wavelength dependence of the irradiance threshold together with tissue optical data was used to estimate the wavelength dependence of the energy threshold at various cutting depths. For focusing depths up to 200 μm, pulse energies required for surgery are smallest for < 800 nm. However, the energy minimum shifts to wavelengths around 1350 nm for z = 500 μm, and to the region around 1700 nm for z = 1 mm.
We developed modeling tools for optical breakdown events in water that span various phases reaching from breakdown initiation via solvated electron generation, through laser induced-plasma formation and temperature evolution in the focal spot to the later phases of cavitation bubble dynamics and shock wave emission and applied them to a large parameter space of pulse durations, wavelengths, and pulse energies.
The rate equation model considers the interplay of linear absorption, photoionization, avalanche ionization and recombination, traces thermalization and temperature evolution during the laser pulse, and portrays the role of thermal ionization that becomes relevant for T > 3000 K. Modeling of free-electron generation includes recent insights on breakdown initiation in water via multiphoton excitation of valence band electrons into a solvated state at Eini = 6.6 eV followed by up-conversion into the conduction band level that is located at 9.5 eV.
The ability of tracing the temperature evolution enabled us to link the model of laser-induced plasma formation with a hydrodynamic model of plasma-induced pressure evolution and phase transitions that, in turn, traces bubble generation and dynamics as well as shock wave emission. This way, the amount of nonlinear energy deposition in transparent dielectrics and the resulting material modifications can be assessed as a function of incident laser energy. The unified model of plasma formation and bubble dynamics yields an excellent agreement with experimental results over the entire range of investigated pulse durations (femtosecond to nanosecond), wavelengths (UV to IR) and pulse energies.
We present a novel experimental setup to intravitally induce and monitor tissue lesions intravitally at a subcellular level
in murine small intestinal mucosa. Using single 355-nm, 500-ps laser pulses coupled to a two-photon microscope, we
induced optical breakdown with subsequent cavitation bubble formation in the tissue. Imaging was based on spectrally
resolved two-photon excited tissue autofluorescence, while online-dosimetry of the induced microbubbles was done by a
cw probe-beam scattering technique. From the scattering signal, the bubble size and dynamics could be deduced on a ns
time scale. In turn, this signal could be used to control the damage size. This was important for reproducible production
of minute effects in the tissue, despite strong biological variations in tissue response to pulsed laser irradiation.
After producing local UV damage, cells appeared dark, probably due to destruction of mitochondria and loss of
NAD(P)H fluorescence. Within 10 min after cell damage, epithelial cells adjacent to the injured area migrated into the
wound to cover the denuded area, resulting in extrusion of the damaged cells from the epithelial layer. Using the nuclear
acid stain propidium iodide, we could show that UV pulses induced cell membrane damage with subsequent necrosis,
rather than apoptosis. For lesions without disruption of the basement membrane, we did not detect migration of immune
cells toward the injured area within observation periods of up to 5 hours.
This model will be used in further studies to investigate the intrinsic repair system and immune response to laserinduced
lesions of intestinal epithelium in vivo.
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