Robust nanopatterning by laser-induced dewetting of metal nanofilms

We have observed nanopattern formation with robust and controllable spatial ordering by laser-induced dewetting in nanoscopic metal films. Pattern evolution in Co film of thickness 1\leq h\leq8 nm on SiO_{2} was achieved under multiple pulse irradiation using a 9 ns pulse laser. Dewetting leads to the formation of cellular patterns which evolve into polygons that eventually break up into nanoparticles with monomodal size distribution and short range ordering in nearest-neighbour spacing R. Spatial ordering was attributed to a hydrodynamic thin film instability and resulted in a predictable variation of R and particle diameter D with h. The length scales R and D were found to be independent of the laser energy. These results suggest that spatially ordered metal nanoparticles can be robustly assembled by laser-induced dewetting

Quantification of optical absorption coefficient from acoustic spectra in the optical diffusive regime using photoacoustic microscopy

Photoacoustic (PA) tomography (PAT) can image optical absorption contrast with ultrasonic spatial resolution in the optical diffusive regime. Multi-wavelength PAT can noninvasively monitor hemoglobin oxygen saturation (sO_2) with high sensitivity and fine spatial resolution. However, accurate quantification in PAT requires knowledge of the optical fluence distribution, acoustic wave attenuation, and detection system bandwidth. We propose a method to circumvent this requirement using acoustic spectra of PA signals acquired at two optical wavelengths. With the acoustic spectral method, the absorption coefficients of an oxygenated bovine blood phantom at 560 and 575 nm were quantified with errors of <5%

In Vivo Photoacoustic Tomography of Chemicals: High-Resolution Functional and Molecular Optical Imaging at New Depths

High-resolution volumetric optical imaging modalities, such as confocal microscopy, two-photon microscopy, and optical coherence tomography, have become increasingly important in the biomedical imaging field. However, due to strong light scattering, the penetration depths of these imaging modalities are limited to the optical transport mean free path in biological tissues, for example, ∼1 mm in the skin. Photoacoustic tomography (PAT), an emerging hybrid imaging modality that can provide strong endogenous and exogenous optical absorption contrasts with high ultrasonic spatial resolution using the photoacoustic (PA) effect, has overcome the fundamental depth limitation. The image resolution is scalable with the ultrasonic frequency. The imaging depth is limited to the reach of photons and up to a few centimeters deep in biological tissues. This Review will focus on the following aspects of PAT described in works published from 2003 to 2009: (1) multiscale PAT systems, (2) morphological and functional PAT using intrinsic contrasts (hemoglobin or melanin), and (3) functional and molecular PAT using exogenous contrast agents (organic dyes, nanoparticles, reporter genes, or fluorescence proteins)

In vivo functional human imaging using photoacoustic microscopy: response to ischemic and thermal stimuli

We report results of two in vivo functional human imaging experiments using photoacoustic microscopy. In Experiment 1, the hemodynamic response to an ischemic event was measured. The palm of a volunteer was imaged and a single cross-section was monitored while periodic arterial occlusions were administered using a blood pressure cuff wrapped around the upper arm and inflated to ~280 mmHg. Significant relative decreases in oxygen saturation (sO_2) and total hemoglobin (HbT) were observed during periods of ischemia. Upon release of the occlusion, significant relative increases in sO_2 and HbT due to post-occlusive reactive hyperemia were recorded. Experiment 2 explored the vascular response to a local, external thermal stimulus. Thermal hyperemia is a common physiological phenomenon and thermoregulation function in which blood flow to the skin is increased to more efficiently exchange heat with the ambient environment. The forearm of a volunteer was imaged and a single cross-section was monitored while the imaged surface was exposed to an elevated temperature of ~46°C. Due to thermal hyperemia, relative increases in sO_2 and HbT were measured as the temperature of the surface was raised. These results may contribute as clinically relevant measures of vascular functioning for detection and assessment of vascular related diseases

Quantitative photoacoustic microscopy of optical absorption coefficients from acoustic spectra in the optical diffusive regime

Photoacoustic (PA) microscopy (PAM) can image optical absorption contrast with ultrasonic spatial resolution in the optical diffusive regime. Conventionally, accurate quantification in PAM requires knowledge of the optical fluence attenuation, acoustic pressure attenuation, and detection bandwidth. We circumvent this requirement by quantifying the optical absorption coefficients from the acoustic spectra of PA signals acquired at multiple optical wavelengths. With the acoustic spectral method, the absorption coefficients of an oxygenated bovine blood phantom at 560, 565, 570, and 575 nm were quantified with errors of <3%. We also quantified the total hemoglobin concentration and hemoglobin oxygen saturation in a live mouse. Compared with the conventional amplitude method, the acoustic spectral method provides greater quantification accuracy in the optical diffusive regime. The limitations of the acoustic spectral method was also discussed

Nonlinear photoacoustic spectroscopy of hemoglobin

As light intensity increases in photoacoustic imaging, the saturation of optical absorption and the temperature dependence of the thermal expansion coefficient result in a measurable nonlinear dependence of the photoacoustic (PA) signal on the excitation pulse fluence. Here, under controlled conditions, we investigate the intensity-dependent photoacoustic signals from oxygenated and deoxygenated hemoglobin at varied optical wavelengths and molecular concentrations. The wavelength and concentration dependencies of the nonlinear PA spectrum are found to be significantly greater in oxygenated hemoglobin than in deoxygenated hemoglobin. These effects are further influenced by the hemoglobin concentration. These nonlinear phenomena provide insights into applications of photoacoustics, such as measurements of average inter-molecular distances on a nm scale or with a tuned selection of wavelengths, a more accurate quantitative PA tomography

Optimal oblique light illumination for photoacoustic microscopy beyond the diffusion limit

To image beyond the quasi-ballistic photon regime, photoacoustic tomography systems must rely on diffuse photons; however, there still exists an optimal illumination pattern that results in the largest number of photons reaching a target at a given depth. Many photoacoustic imaging systems incorporate weak optical focusing through oblique or dark-field illumination, but these systems are not often optimized for deep light delivery. Multiple parameters and constraints, particularly for in vivo imaging, need to be considered to determine the optimal illumination scheme for a given system: beam diameter, incident angle, pulse repetition rate, laser fluence, and target depth. Monte Carlo simulations of varied beam geometries and incident angles show the best optical illumination schemes for different imaging depths. Further an analytic model based on the diffusion theory provides a rapid method of determining the optimal beam size and incident angle for a given target depth and agrees well with the simulations. The results reveal the most efficient optical focal position to maximize the number of photons delivered to a target depth, therein maximizing the PA signal. The principles and results discussed here are not limited to the system investigated, but can be applied to other system configurations to improve the photoacoustic signal strength

In vivo multiscale photoacoustic microscopy of human skin

Scalability is a key feature of photoacoustic microscopy (PAM). Reports have shown that PAM systems can be designed to possess sub-micron resolution at shallow depths or penetrate centimeters deep at the expense of resolution while the number of resolved pixels in the depth direction remains high. This capability to readily tune the imaging parameters while maintaining the same inherent contrast could be extremely useful for a variety of biomedical applications. Human skin, with its layered vascular structure whose dimensions scale with depth, provides an ideal imaging target to illustrate this advantage. Here, we present results from in vivo human skin imaging experiments using two different PAM systems, an approach which enables better characterization of the cutaneous microvasculature throughout the imaging depth. Specifically, we show images from several distinct areas of skin: the palm and the forearm. For each region, the same area was imaged with both an optical-resolution PAM (OR-PAM) and an acoustic-resolution PAM (AR-PAM), and the subsequent images were combined into composite images. The OR-PAM provides less than 5 μm lateral resolution, capable of imaging the smallest capillary vessels, while the AR-PAM enables imaging at depths of several millimeters. Several structures are identifiable in the ORPAM images which cannot be differentiated in AR-PAM images, namely thin epidermal and stratum corneum layers, undulations in the dermal papillae, and capillary loops. However, the AR-PAM provides images of larger vessels, deeper than the OR-PAM can penetrate. These results demonstrate how PAM's scalability can be utilized to more fully characterize cutaneous vasculature, potentially impacting the assessment of numerous cardiovascular related and cutaneous diseases

Functional photoacoustic microscopy of pH

pH is a tightly regulated indicator of metabolic activity. In mammalian systems, imbalance of pH regulation may result from or result in serious illness. Even though the regulation system of pH is very robust, tissue pH can be altered in many diseases such as cancer, osteoporosis and diabetes mellitus. Traditional high-resolution optical imaging techniques, such as confocal microscopy, routinely image pH in cells and tissues using pH sensitive fluorescent dyes, which change their fluorescence properties with the surrounding pH. Since strong optical scattering in biological tissue blurs images at greater depths, high-resolution pH imaging is limited to penetration depths of 1mm. Here, we report photoacoustic microscopy (PAM) of commercially available pH-sensitive fluorescent dye in tissue phantoms. Using both opticalresolution photoacoustic microscopy (OR-PAM), and acoustic resolution photoacoustic microscopy (AR-PAM), we explored the possibility of recovering the pH values in tissue phantoms. In this paper, we demonstrate that PAM was capable of recovering pH values up to a depth of 2 mm, greater than possible with other forms of optical microscopy