Fluorescent lifetime imaging is an optical technique that facilitates imaging molecular interactions and cellular functions. Because the excited lifetime of a fluorophore is sensitive to its local microenvironment,<sup>1, 2 </sup>measurement of fluorescent lifetimes can be used to accurately detect regional changes in temperature, pH, and ion concentration. However, typical state of the art fluorescent lifetime methods are severely limited when it comes to acquisition time (on the order of seconds to minutes) and video rate imaging. Here we show that compressed ultrafast photography (CUP) can be used in conjunction with fluorescent lifetime imaging to overcome these acquisition rate limitations. Frame rates up to one hundred billion frames per second have been demonstrated with compressed ultrafast photography using a streak camera.<sup>3</sup> These rates are achieved by encoding time in the spatial direction with a pseudo-random binary pattern. The time domain information is then reconstructed using a compressed sensing algorithm, resulting in a cube of data (x,y,t) for each readout image. Thus, application of compressed ultrafast photography will allow us to acquire an entire fluorescent lifetime image with a single laser pulse. Using a streak camera with a high-speed CMOS camera, acquisition rates of 100 frames per second can be achieved, which will significantly enhance our ability to quantitatively measure complex biological events with high spatial and temporal resolution. In particular, we will demonstrate the ability of this technique to do single-shot fluorescent lifetime imaging of cells and microspheres.
Wide-field microscopy, where full images are obtained simultaneously, is limited by the power available from speckle-free light sources. Currently, the vast majority of wide-field microscopes use either mercury arc lamps, or LEDs as the illumination source. The power available from these sources limits wide-field fluorescent microscopy to tens of microseconds temporal resolution. Lasers, while capable of producing high power and short pulses, have high spatial coherence. This leads to the formation of laser speckle that makes such sources unsuitable for wide-field imaging applications. Random Raman lasers offer the best of both worlds by producing laser-like intensities, short, nanosecond-scale, pulses, and low spatial coherence, speckle-free, output. These qualities combine to make random Raman lasers 4 orders of magnitude brighter than traditional wide-field microscopy light sources. Furthermore, the unique properties of random Raman lasers make possible the entirely new possibilities of wide-field fluorescence lifetime imaging or wide-field Raman microscopy. We will introduce the relevant physics that give rise to the unique properties of random Raman lasing, and demonstrate early proof of principle results demonstrating random Raman lasing emission being used as an imaging light source. Finally, we will discuss future directions and elucidate the benefits of using random Raman lasers as a wide-field microscopy light source.
Monte Carlo simulations are widely considered to be the gold standard for studying the propagation of light in turbid media. However, due to the probabilistic nature of these simulations, large numbers of photons are often required in order to generate relevant results. Here, we present methods for reduction in the variance of dose distribution in a computational volume. Dose distribution is computed via tracing of a large number of rays, and tracking the absorption and scattering of the rays within discrete voxels that comprise the volume. Variance reduction is shown here using quasi-random sampling, interaction forcing for weakly scattering media, and dose smoothing via bi-lateral filtering. These methods, along with the corresponding performance enhancements are detailed here.
Monte Carlo simulations are widely considered to be the gold standard for studying the propagation of light in turbid media. However, traditional Monte Carlo methods fail to account for diffraction because they treat light as a particle. This results in converging beams focusing to a point instead of a diffraction limited spot, greatly effecting the accuracy of Monte Carlo simulations near the focal plane. Here, we present a technique capable of simulating a focusing beam in accordance to the rules of Gaussian optics, resulting in a diffraction limited focal spot. This technique can be easily implemented into any traditional Monte Carlo simulation allowing existing models to be converted to include accurate focusing geometries with minimal effort. We will present results for a focusing beam in a layered tissue model, demonstrating that for different scenarios the region of highest intensity, thus the greatest heating, can change from the surface to the focus. The ability to simulate accurate focusing geometries will greatly enhance the usefulness of Monte Carlo for countless applications, including studying laser tissue interactions in medical applications and light propagation through turbid media.
Anderson localization, also known as strong localization, is the absence of diffusion in turbid media resulting from wave interference. The effect was originally predicted for electron motion, and is widely known to exist in systems of less than 3 dimensions. However, Anderson localization of optical photons in 3 dimensional systems remains an elusive and controversial topic. Random Raman lasing offers the unique combination of large gain and virtually zero absorption. The lack of absorption makes long path length, localized modes preferred. The presence of gain offsets what little absorption is present, and preferentially amplifies localized modes due to their large Q factors compared with typical low Q modes present in complex media. Random Raman lasers exhibit several experimentally measured properties that diverge from classical, particle-like, diffusion. First, the temporal width of the emission being 1 to a few nanoseconds in duration when it is pumped with a 50 ps laser is a full order of magnitude longer than is predicted by Monte Carlo simulations. Second, the random Raman laser emission is highly multi-mode, consisting of hundreds of simultaneous lasing modes. This is in contrast to early theoretical results and back of the envelope arguments that both suggest that only a few modes should be present. We will present the evidence that suggests a divergence from classical diffusion theory. One likely explanation, that is consistent with all of these anomalies, is the presence of high-Q localized modes consistent with Anderson localization.
Contamination of the water source and air pollution are two major problems that must be faced in the coming years. The increasing worldwide contamination of freshwater systems with thousands of industrial and natural chemical compounds is one of the key environmental problems facing humanity today. It is estimated that pathogens in water cause more than 2 million deaths annually. Additionally, traditional water quality assessment methods, such as liquid chromatography and mass spectroscopy, are expensive and time consuming from sample collection to analysis. Low cost tools are needed which can provide high sensitivity in sensing, while remaining portable and providing near real time analysis. Here, we present a low cost integrating cavity that can be used for highly sensitive environmental sensing.
The time-temperature effects of laser radiation exposure are investigated as a function of wavelength. We experimentally measure the thermal response of tissue to laser radiation ranging in wavelength from 1100 nm to 1550 nm. Simulations were then performed to estimate damage thresholds.
The time-temperature effects of laser radiation exposure are investigated as a function of wavelength. Here, we report the thermal response of bulk tissue as a function of wavelength from 700 to 1064 nm. Additionally, Monte Carlo simulations were used to verify the thermal response measured and predict damage thresholds based on the response.
Raman spectroscopy is a powerful technique that can be used to obtain detailed chemical information about a
system without the need for chemical markers. It has been widely used for a variety of applications such as
cancer diagnosis and material characterization. However, Raman scattering is a highly inefficient process,
where only one in 10<sup>11</sup> scattered photons carry the needed information. Several methods have been developed
to enhance this inherently weak effect, including surface enhanced Raman scattering and coherent anti-Stokes
Raman scattering. These techniques suffer from drawbacks limiting their commercial use, such as the need
for spatial localization of target molecules to a ‘hot spot’, or the need for complex laser systems.
Here, we present a simple instrument to enhance spontaneous Raman scattering using elastic light scattering.
Elastic scattering is used to substantially increase the interaction volume. Provided that the scattering
medium exhibits very low absorption in the spectral range of interest, a large enhancement factor can be
attained in a simple and inexpensive setting. In our experiments, we demonstrate an enhancement of 10<sup>7</sup> in
Raman signal intensity. The proposed novel device is equally applicable for analyzing solids, liquids, and
Tuberculosis, caused by <i>Mycobacterium tuberculosis (Mtb)</i>, currently affects roughly one-third of the world’s population. Drug resistant strains of Mtb decrease the effectiveness of current therapeutics and demand the development of new antimicrobial therapies. In addition, the current vaccine, Bacille Calmette Guérin (BCG), has variable efficacy for disease prevention in different populations. Animal studies are often limited by the need to sacrifice at discrete time points for pathology and tissue homogenization, which greatly reduces spatial and temporal resolution. Optical imaging offers the potential for a minimally-invasive solution to imaging on a macroscopic and microscopic scale, allowing for high resolution study of infection. We have integrated a fluorescence microendoscope into a whole-animal optical imaging system, allowing for simultaneous microscopic and macroscopic imaging of tdTomato expressing BCG <i>in vivo</i>. A 535 nm LED was collimated and launched into a 10,000 element fiber bundle with an outer diameter of 0.66 mm. The fiber bundle can be inserted through an intra-tracheal catheter into the lung of a mouse. Fluorescence emission can either be (1) collected by the bundle and imaged onto the surface of a CCD camera for localized detection or (2) the fluorescence can be imaged by the whole animal imaging system providing macroscopic information. Results from internal localized excitation and external whole body detection indicate the potential for imaging bacterial infections down to 100 colony forming units. This novel imaging technique has the potential to allow for functional studies, enhancing the ability to assess new therapeutic agents.