Paul J. D. Whiteside

Photoacoustic spectroscopy of β-hematin.

Malaria affects over 200 million individuals annually, resulting in 800,000 fatalities. Current tests use blood smears and can only detect the disease when 0.1-1% of blood cells are infected. We are investigating the use of photoacoustic flowmetry to sense as few as one infected cell among 10 million or more normal blood cells, thus diagnosing infection before patients become symptomatic. Photoacoustic flowmetry is similar to conventional flow cytometry, except that rare cells are targeted by nanosecond laser pulses to induce ultrasonic responses. This system has been used to detect single melanoma cells in 10 ml of blood. Our objective is to apply photoacoustic flowmetry to detection of the malaria pigment hemozoin, which is a byproduct of parasite-digested hemoglobin in the blood. However, hemozoin is difficult to purify in quantities greater than a milligram, so a synthetic analog, known as β-hematin was derived from porcine haemin. The specific purpose of this study is to establish the efficacy of using β-hematin, rather than hemozoin, for photoacoustic measurements. We characterized β-hematin using UV-vis spectroscopy, TEM, and FTIR, then tested the effects of laser irradiation on the synthetic product. We finally determined its absorption spectrum using photoacoustic excitation. UV-vis spectroscopy verified that β-hematin was distinctly different from its precursor. TEM analysis confirmed its previously established nanorod shape, and comparison of the FTIR results with published spectroscopy data showed that our product had the distinctive absorbance peaks at 1661 and 1206 cm(-1). Also, our research indicated that prolonged irradiation dramatically alters the physical and optical properties of the β-hematin, resulting in increased absorption at shorter wavelengths. Nevertheless, the photoacoustic absorption spectrum mimicked that generated by UV-vis spectroscopy, which confirms the accuracy of the photoacoustic method and strongly suggests that photoacoustic flowmetry may be used as a tool for diagnosis of malaria infection.

Nanoparticle Mediated Thermal Ablation of Breast Cancer Cells Using a Nanosecond Pulsed Electric Field

In the past, ablation of cancer cells using radiofrequency heating techniques has been demonstrated, but the current methodology has many flaws, including inconsistent tumor ablation and significant ablation of normal cells. Other researchers have begun to develop a treatment that is more selective for cancer cells using metallic nanoparticles and constant electric field exposure. In these studies, cell necrosis is induced by heating antibody functionalized metallic nanoparticles attached to cancer cells. Our approach to studying this phenomenon is to use similarly functionalized metallic nanoparticles that are specific for the T47D breast cancer cell line, exposing these nanoparticle cell conjugates to a nanosecond pulsed electric field. Using fluorescent, polystyrene-coated, iron-oxide nanoparticles, the results of our pilot study indicated that we were able to ablate up to approximately 80% of the cells using 60 ns pulses in increasing numbers of pulses and up to approximately 90% of the cells using 300 ns pulses in increasing numbers of pulses. These quantities of ablated cells were achieved using a cumulative exposure time 6 orders of magnitude less than most in vitro constant electric field studies.

Photoacoustic measurement of refractive index of dye solutions and myoglobin for biosensing applications

Current methods of determining the refractive index of chemicals and materials, such as ellipsometry and reflectometry, are limited by their inability to analyze highly absorbing or highly transparent materials, as well as the required prior knowledge of the sample thickness and estimated refractive index. Here, we present a method of determining the refractive index of solutions using the photoacoustic effect. We show that a photoacoustic refractometer can analyze highly absorbing dye samples to within 0.006 refractive index units of a handheld optical refractometer. Further, we use myoglobin, an early non-invasive biomarker for malignant hyperthermia, as a proof of concept that this technique is applicable for use as a medical diagnostic. Comparison of the speed, cost, simplicity, and accuracy of the techniques shows that this photoacoustic method is well-suited for optically complex systems.

Total internal reflection photoacoustic spectroscopy for the detection of β-hematin.

Evanescent field sensing methods are currently used to detect many different types of disease markers and biologically important chemicals such as the HER2 breast cancer receptor. Hinoue et al. used Total Internal Reflection Photoacoustic Spectroscopy (TIRPAS) as a method of using the evanescent field to detect an optically opaque dye at a sample interface. Although their methods were successful at detecting dyes, the results at that time did not show a very practical spectroscopic technique, which was due to the less than typical sensitivity of TIRPAS as a spectroscopy modality given the low power (≈ 1 to 2 W) lasers being used. Contrarily, we have used an Nd:YAG laser with a five nanosecond pulse that gives peak power of 1 MW coupled with the TIRPAS system to increase the sensitivity of this technique for biological material sensing. All efforts were focused on the eventual detection of the optically absorbing material, hemozoin, which is created as a byproduct of a malarial infection in blood. We used an optically analogous material, β-hematin, to determine the potential for detection in the TIRPAS system. In addition, four properties which control the sensitivity were investigated to increase understanding about the sensors function as a biosensing method.

Total internal reflection photoacoustic detection spectroscopy

Total Internal Reflection Photoacoustic Spectroscopy (TIRPAS) is a method that exploits the evanescent field of a nanosecond duration laser pulse reflecting off a glass/water interface to generate photoacoustic responses. These photoacoustic events are generated in light absorbing analytes suspended in the fluid medium in contact with the glass that are within the penetration depth of the evanescent wave. This method has been employed in previous studies by Hinoue et al. Hinoue et al. used an optically chopped HeNe laser at 632.8 nm to detect Brilliant Blue FCF dye at different angles of incidence. In recent years, the advent of high power nanosecond pulsed tunable lasers has allowed for the re-visitation of the TIRPAS idea under stress confinement and orders of magnitude larger peak energy conditions. Compared to conventional detection methods, this approach has the potential to detect much smaller quantities of disease indicators, such as circulating tumor cells and hemazoin crystals in malaria, than other optical methods. The detection limit of the TIRPAS system was quantified using chlorazol black solution with an absorption coefficient of 55 cm-1 at 532 nm. Interaction with the evanescent field was verified by varying the angle of incidence of the probe laser beam that generated the photoacoustic waves, thereby changing the penetration depth of the evanescent field as well as the photoacoustic spectroscopy effect from angled excitation.

Planar waveguide light transmission modality for backward-mode photoacoustic tomography

Prior research in photoacoustic tomography has consistently demonstrated its ability to image structures near the surface of tissue with a high degree of optical contrast. However, despite significant advancements in the field, there has been little to no development of clinical applications for photoacoustic tomography, principally due to the requirement for backwardmode operation, i.e., it must detect the photoacoustic signal on the same side of the tissue as the incident laser light. This results in the standard ultrasonic transducer occluding the path of the inciting laser beam. Therefore, developing a technique to deliver light into the tissue, while incorporating commonly available ultrasonic detection equipment without occluding the beam propagation or modifying the equipment in any way, would provide a significant benefit to the field, and potentially improve its clinical applicability. Here, we propose a new method to accomplish this aim, using planar optical waveguides that employ the optical tunneling phenomenon to transmit light directly into tissue (pig skin) through physical contact with the sample. A commercially available, 10MHz, unfocused ultrasonic transducer was positioned on the rear face of the waveguide and was used to detect photoacoustic signals generated within the tissue as the signals propagated perpendicularly through the waveguide substrate. Unlike alternative solutions to the occlusion problem, this modality does not necessitate the use of custom manufactured transducers, expensive dichroics, or additional laser systems, and thereby represents a viable approach for the easy implementation of photoacoustic tomography in a clinical setting.

Metal-clad waveguide characterization for contact-based light transmission into tissue

As contemporary laser dermatology procedures, like tattoo removal and skin resurfacing, become more popular, the complications of their operation are also becoming more prevalent. Frequent incidences of over-exposure, ocular injury, and excessive thermal damage represent mounting concerns for those seeking such procedures; moreover, each of these problems is a direct consequence of the standard, free-space method of laser transmission predominantly used in clinical settings. Therefore, an alternative method of light transmission is needed to minimize these problems. Here, we demonstrate and characterize an alternative method that uses planar waveguides to deliver light into sample tissue via direct contact. To do this, slab substrates made from glass were clad in layers of titanium and silver, constraining the light within the waveguide along the waveguide’s length. By creating active areas on the waveguide surface, the propagating light could then optically tunnel into the tissue sample, when the waveguide was brought into contact with the tissue. SEM and EDS were used to characterize the metal film thickness and deposition rates onto the glass substrates. Laser light from a Q-switched Nd:YAG source operating at 532nm was coupled into the waveguide and transmitted into samples of pig skin. The amount of light transmitted was measured using photoacoustics techniques, in conjunction with a photodiode and integrating sphere. Transmitting light into tissue in this manner effectively resolves or circumvents the complications caused by free-space propagation methods as it reduces the operating distance to 0, which prevents hazardous back-reflections and allows for the ready incorporation of contact cooling technologies.

CASE STUDY OF HIGH BLOOD GLUCOSE CONCENTRATION EFFECTS OF 850 MHZ ELECTROMAGNETIC FIELDS USING GTEM CELL

The efiect of 850MHz electromagnetic radiation on diabetic blood at 2W and 60W power levels was investigated and compared with normal blood cells. The power levels respectively represent radiations from a cell phone and the cell phone tower, both operating 850MHz. A GTEM cell was designed for the tests to generate the desired uniform electromagnetic fleld and power in a shielded environment. Blood samples, having normal and high glucose concentrations, were placed in the usable area inside the GTEM cell for 10, 30, 60minutes and the glucose levels and red and white blood cell viabilities were monitored and compared with the controls. Results show that the 850MHz exposure signiflcantly in∞uences the blood cell counts and the glucose level in both normal and high glucose blood samples. In cell survivability analysis in normal blood samples it was found that the white blood cells are signiflcantly higher than the control at 60min exposure from cell phone radiation, while both the white and red blood cell are signiflcantly higher following a 30min exposure from tower radiation. For high glucose blood tests at 30 and 60min exposure times, the tower radiation for 60min and the cell phone radiation at both the exposure times show signiflcantly changes in white blood cell counts, whereas there was no efiect in red blood cells. Also, for 30 and 60min exposure times, the glucose

Ultrasonic modulation of tissue optical properties in ex vivo porcine skin to improve transmitted transdermal laser intensity

Applications of light‐based energy devices involving optical targets within the dermis frequently experience negative side‐effects resultant from surface scattering and excess optical absorption by epidermal melanin. As a broadband optical absorber, melanin decreases the efficacy of light‐based treatments throughout the ultraviolet, visible, and near‐infrared spectra while also generating additional heat within the surface tissue that can lead to inflammation or tissue damage. Consequently, procedures may be performed using greater energy densities to ensure that the target receives a clinically relevant dose of light; however, such practices are limited, as doing so tends to exacerbate the detrimental complications resulting from melanin absorption of treatment light. The technique presented herein represents an alternative method of operation aimed at increasing epidermal energy fluence while mitigating excess absorption by unintended chromophores. The approach involves the application of continuously pulsed ultrasound to modulate the tissues optical properties and thereby improve light transmission through the epidermis.

Response of high blood glucose level to GTEM cell electromagnetic fields to simulate cell phone radiation

Summary form only given. Due to the proliferation of devices emitting electromagnetic radiation, there has been a renewed focus to study the harmful, as well as the beneficial effects of electromagnetic radiation. Radiation from cell phone and devices operating in GSM-900 band are likely to have effects on living organism and needs investigation. Recent works in this area include the effects of cellular phone radiation on blood-brain barrier connections1, cell toxicity2 and on carcinogenic3. In diabetic related studies, which are an important topic area, studies on the effect of cell phone exposure on brain glucose metabolism4 and the effects of transient EM fields on the plasma glucose levels in Type 1 and Type 2 diabetics5 have been reported. The study of the effects of cell phone radiation to blood sugar levels, therefore, is another area of interest and needs investigation.