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Laser-based optical control technologies have shown promise in achieving high spatiotemporal precision. However, existing methods face challenges in real-time target selection and manipulation. To overcome these limitations, we present a real-time precision opto-control (RPOC) technology, which is a closed-loop optoelectronic system that is built upon a laser scanning confocal fluorescence microscope and integrates chemical-specific optical detection, real-time decision-making, and precise optical manipulation at target sites. Using RPOC, we demonstrated precise inducing reactive oxygen species (ROS) solely at selected targets and monitoring ROS-induced changes in microtubule polymerization dynamics. We also selectively inhibit tubulin polymerization using RPOC paired with a photoswitchable inhibitor.
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Nanosecond electric pulses (nsEP) are effective in biomedical applications like cancer treatment, gene therapy, and drug delivery due to their ability to influence cellular membranes and intracellular processes without thermal damage. However, the high electric fields required for these bioeffects present challenges. Recent studies suggest that MHz compression of nsEP could lead to similar effects at lower field strengths, enhancing safety and efficacy. This study leverages streak camera and optical streaking microscopy to examine membrane charging dynamics from nsEP bursts. Results will broaden understanding of membrane responses to nsEP, potentially improving their effectiveness and safety in biomedical applications.
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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.
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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.
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We demonstrate precise cellular laser ablation on SH-SY5Y and onion cells by using an advanced 1.95μm nanosecond-pulsed thulium-doped fibre laser (TDFL). The TDFL offers a high degree of control on pulse parameters, which enables good thermal and mechanical confinement during the laser ablation and results in a high precision of 30μm, with minimal carbonisation or collateral damage to surrounding cells. The realisation of precise cellular ablation from a TDFL will open up new applications in microsurgery for disease treatment, benefitting patients and researchers worldwide.
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The successful laser ablation of clinically relevant tissue models by means of picosecond laser pulses is presented. This is a potential alternative to overcoming limitations of conventional tumour-surgery tools in terms of precision and thermal damage. The correlation of high-speed imaging of the ablation process, schlieren imaging of the resulting plume dynamics and a histopathological analysis of the post-process tissue morphology enables optimisation of the tissue removal rate whilst avoiding adverse cavitation effects. This facilitates minimal collateral thermal damage. Effective tissue removal is presented for the epithelial laser ablation of colonic tissue; with translation of this process towards infiltrating brain and head and neck cancer surgery further discussed.
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Novel Applications of Lasers and Light in Biomedicine I
Until today, the role of lasers in surgery has been mostly limited to tissue cutting (laser scalpels), which roboticists have supported by developing micro-mechanical systems for accurate, tremor-free laser aiming. As medical science evolves, we keep discovering new ways in which laser light can be used for surgical treatment: There are new types of laser-based treatment being pioneered where thermal necrosis – not resection! – is the goal. For these treatments to work, besides laser aiming, it is vital to also monitor and control the interactions between the laser and the tissue. These interactions, however, are notoriously hard to control, both by humans and machines, as they involve fast, highly nonlinear physical phenomena that can be challenging to model and even perceive adequately. My research vision is to enable a new generation of surgical robots, capable of intelligently monitoring and controlling surgical laser-tissue interactions. These robots will continuously monitor the status of a procedure and assist physicians in regulating laser delivery to achieve the desired clinical outcomes.
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Cellular photomodulation holds promising potential in biomedical research, However, this technique is typically performed manually at low speed. We have successfully developed an integrated femtosecond laser (fsL) cell stimulation system based on a constant radius lab-on-a-disc (LOAD) platform in which a concentric microchannel for rearranging cells to a monolayer under centrifugal force can be engraved in a polymethyl methacrylate layer in situ while the disc is in spinning mode. The system can perform multi-photon cell stimulation, which subsequently leads to molecular signaling modulation of cells in a high-throughput and highly automatic manner.
This project was supported by General Research Fund (GRF) : 14204621, 14207920 and 14207419.
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Novel Applications of Lasers and Light in Biomedicine II
A photonics localization method, called inverse participation ratio (IPR), is adeptly applied to elucidate the effects of probiotics and alcohol on colon cancer by quantifying the DNA molecular-specific spatial structural changes in colon cancer cell nuclei on a colon cancer mouse model via confocal imaging. The IPR light localization technique measures the degree of structural disorder of DNA molecular-specific spatial mass density fluctuations. The nuclear structural alterations in colon cancer cell nuclei have been known to begin at the nano-to-submicron level, which precedes and predicts more prominent microscopic observations later in the disease. The effects of probiotics on alcohol-treated colon cancer are not a well-understood problem. However, probiotics like Lactobacillus have proven effective in enhancing colon cell/tissue functions. The IPR study results show that alcohol treatment enhances colon cancer, and the treatment of probiotics on alcohol-treated colon cancer tries to bring colon cancer less severe to normal. We acknowledge the grant NIH- R21CA260147.
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Numerical Approaches Simulating Laser-Tissue Interactions and Response
We combine continuous-wave (CW) and time-resolved (TR) near-infrared spectroscopy (NIRS) to effectively quantify tissue optical properties (OPs) of both shallow and deep regions in human heads. Utilizing iterative curve fitting in which artificial neural networks trained with Monte-Carlo simulated data serve as the forward model, we derive the OPs of interest such as the gray matter through TR NIRS with the assumption of known OPs of shallower tissue regions estimated from broadband CW-NIRS measurements. Preliminary results indicate improved accuracy in quantifying deep tissue absorption coefficients, while presenting more challenges in estimating scattering coefficients. Further research and refinements are essential to address this challenge to facilitate a more thorough understanding of photon propagation in the head composed of complex biological tissues.
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Photothermal, Photochemical, Photo-Oxidative, and Photomechanical Interactions
Ultrashort pulse lasers are currently used in a variety of applications, including laser eye surgery. However, laser safety standards do not consider the potential hazard to the eye due to the nonlinear interaction of ultrashort lasers with the ocular tissues. We used a single NIR femtosecond pulse to determine the peak pulse energies that generated a supercontinuum within the eye of anesthetized porcine subjects and resulted in retinal alterations. The results of this study inform the laser safety standards about hazards to the eye due to the supercontinuum generated by nonlinear effects in the aqueous media of the eye.
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High-dose laser exposure to tissue causes thermal damage and significant changes in tissue optical properties. Samples of porcine dermis and subcutaneous fat were immersed in a temperature-controlled water bath to induce a range of thermal damage. Temperature history was recorded to quantify the damage with the Arrhenius integral. Samples were then measured in a double integrating sphere setup and optical coefficients computed using the inverse adding doubling method. The tissues demonstrate non-monotonic changes in optical properties with respect to induced thermal damage. These results will inform medical scenarios and computational models where optical interaction with damaged tissues is expected.
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Photo-mediate ultrasound therapy (PUT) is a new anti-vascular technique based on cavitation induced spallation, applies nanosecond laser pulses and ultrasound bursts simultaneously to promote cavitation activity to remove the blood vessel. However, the real time detection of cavitation to guide PUT in real time is still challenging. To better understand the spatial-temporal distribution of cavitation bubbles, we proposed to integrate Doppler optical coherence tomography (DOCT) combined PUT, where the DOCT was used to visualize shockwave-induced cavitation and monitor treatment response in real time. The addition of DOCT to PUT allows for quantitative prescreening and real time monitoring during treatment response, which can improve the safety and effectiveness of the therapy.
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The shortwave infrared (SWIR) optical window has distinct advantages in terms of improved imaging penetration depth and unique optical contrasts in tissue. Here I will describe two new diffuse optical technologies our research team is developing and extending to the SWIR wavelength band: spatial frequency domain imaging (SFDI) and frequency domain diffuse optical spectroscopy (FD-DOS). I will describe the advantages of these techniques and show clinical data related to applications in which these techniques have been deployed: non-invasive blood lipids monitoring and kidney dialysis monitoring. I will also discuss openSFDI, an open hardware project that provides instructions for groups looking to develop their own SFDI systems.
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Cell growth is very important for the development and maintenance of organisms, and it is essential to study factors that affect cell growth, such as gas concentrations that affect cell metabolism, especially oxygen and carbon dioxide. In this study, the dynamic activity of cells according to oxygen concentration was analyzed using a dynamic full-field optical coherence tomography imaging system in a gas-controlled chamber for label-free analysis of living cells. This study aims to understand the relationship between gas supply levels and intracellular activity through non-invasive observation. This discovery could have important implications for biomedicine and biotechnology.
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Visualization-based monitoring provides an empirical and minimally-invasive means for evaluating monolayer cell cultures, but does not translate to three-dimensional microcarrier-based cultures. In toto visualization of cell density and morphology is imperative for producing high quality, high yield cell cultures. Here, the optical properties of commercial polystyrene and custom-fabricated hydrogel microcarriers are compared for compatibility with light-sheet imaging for visualization and enumeration of adherent cells. Additionally, Mie scattering simulations were performed to describe the angular scattering intensity distributions. This study shows the custom hydrogel microcarrier is compatible with in toto non-destructive and non-invasive visualization and monitoring of 3D adherent cell cultures.
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