Thin films continue to show great promise for improving a wide variety of devices in applications such as medical instrumentation, material processing, and astronomical instrumentation. While ellipsometry and reflectometry are standard characterization techniques for determining thickness and refractive index, these techniques tend to require highly reflective or polished films and rely on empirical equations. We have created Quantum Tunneling Photoacoustic Spectroscopy (QTPAS) that uses light induced ultrasound to obtain thickness and refractive index estimates of transparent films. We present QTPAS to be used for the estimation of properties of single layer films as an alternative to ellipsometry and give qualitative sample measurements of the technique's estimated parameters.
Due to the often extreme energies employed, contemporary methods of laser delivery utilized in clinical dermatology allow
for a dangerous amount of high-intensity laser light to reflect off a multitude of surfaces, including the patient’s own skin.
Such techniques consistently represent a clear and present threat to both patients and practitioners alike. The intention of
this work was therefore to develop a technique that mitigates this problem by coupling the light directly into the tissue
via physical contact with an optical waveguide. In this manner, planar waveguides cladded in silver with thin-film active
areas were used to illuminate agar tissue phantoms with nanosecond-pulsed laser light at 532nm. The light then either
refracted or optically tunneled through the active area, photoacoustically generating ultrasonic waves within the phantom,
whose peak-to-peak intensity directly correlated to the internal reflection angle of the beam. Consequently, angular spectra
for energy delivery were recorded for sub-wavelength silver and titanium films of variable thickness. Optimal energy
delivery was achieved for internal reflection angles ranging from 43 to 50 degrees, depending on the active area and
thin film geometries, with titanium films consistently delivering more energy across the entire angular spectrum due to
their relatively high refractive index. The technique demonstrated herein therefore not only represents a viable method of
energy delivery for biological tissue while minimizing the possibility for stray light, but also demonstrates the possibility
for utilizing thin films of high refractive index metals to redirect light out of an optical waveguide.
According to the CDC, breast cancer is the most common cancer and the second leading cause of cancer related
deaths among women. Metastasis, or the presence of secondary tumors caused by the spread of cancer cells via
the circulatory or lymphatic systems, significantly worsens the prognosis of any breast cancer patient. In this
study, a technique is developed to detect circulating breast cancer cells in human blood using a photoacoustic flow
cytometry method. A Q-switched laser with a 5 ns pulse at 532 nm is used to interrogate thousands of cells with
one pulse as they flow through the beam path. Cells which are pigmented, either naturally or artificially, emit an
ultrasound wave as a result of the photoacoustic (PA) effect. Breast cancer cells are targeted with chromophores
through immunochemistry in order to provide pigment. After which, the device is calibrated to demonstrate a
single-cell detection limit. Cultured breast cancer cells are added to whole blood to reach a biologically relevant
concentration of about 25-45 breast cancer cells per 1 mL of blood. An in vitro photoacoustic flow cytometer is
used to detect and isolate these cells followed by capture with the use of a micromanipulator. This method can
not only be used to determine the disease state of the patient and the response to therapy, it can also be used
for genetic testing and in vitro drug trials since the circulating cell can be captured and studied.
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 sensor's function as a biosensing method.
Melanoma is the deadliest form of skin cancer, yet current diagnostic methods are unable to detect early onset of metastatic disease. Patients must wait until macroscopic secondary tumors form before malignancy can be diagnosed and treatment prescribed. Detection of cells that have broken off the original tumor and travel through the blood or lymph system can provide data for diagnosing and monitoring metastatic disease. By irradiating enriched blood samples spiked with cultured melanoma cells with nanosecond duration laser light, we induced photoacoustic responses in the pigmented cells. Thus, we can detect and enumerate melanoma cells in blood samples to demonstrate a paradigm for a photoacoustic flow cytometer. Furthermore, we capture the melanoma cells using microfluidic two phase flow, a technique that separates a continuous flow into alternating microslugs of air and blood cell suspension. Each slug of blood cells is tested for the presence of melanoma. Slugs that are positive for melanoma, indicated by photoacoustic waves, are separated from the cytometer for further purification and isolation of the melanoma cell. In this paper, we evaluate the two phase photoacoustic flow cytometer for its ability to detect and capture metastastic melanoma cells in blood.
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<sup>-1</sup> 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.
Melanoma is the deadliest form of skin cancer, yet current diagnostic methods are inadequately sensitive. Patients
must wait until secondary tumors form before malignancy can be diagnosed and treatment prescribed. Detection of
cells that have broken off the original tumor and flow through the blood or lymph system can provide data for
diagnosing and monitoring cancer. Our group utilizes the photoacoustic effect to detect metastatic melanoma cells,
which contain the pigmented granule melanin. As a rapid laser pulse irradiates melanoma, the melanin undergoes
thermo-elastic expansion and ultimately creates a photoacoustic wave. Thus, melanoma patient's blood samples can
be enriched, leaving the melanoma in a white blood cell (WBC) suspension. Irradiated melanoma cells produce
photoacoustic waves, which are detected with a piezoelectric transducer, while the optically transparent WBCs
create no signals. Here we report an isolation scheme utilizing two-phase flow to separate detected melanoma from
the suspension. By introducing two immiscible fluids through a t-junction into one flow path, the analytes are
compartmentalized. Therefore, the slug in which the melanoma cell is located can be identified and extracted from
the system. Two-phase immiscible flow is a label free technique, and could be used for other types of pathological
Photoacoustic flowmetry has been used to detect single circulating melanoma cells in vitro. Circulating melanoma cells
are those cells that travel in the blood and lymph systems to create secondary tumors and are the hallmark of metastasis.
This technique involves taking blood samples from patients, separating the white blood and melanoma cells from whole
blood and irradiating them with a pulsed laser in a flowmetry set up. Rapid, visible wavelength laser pulses on the order
of 5 ns can induce photoacoustic waves in melanoma cells due to their melanin content, while surrounding white blood
cells remain acoustically passive. We have developed a system that identifies rare melanoma cells and captures them in
50 microliter volumes using suction applied near the photoacoustic detection chamber. The 50 microliter sample is then
diluted and the experiment is repeated using the new sample until only a melanoma cell remains. We have tested this
system on dyed microspheres ranging in size from 300 to 500 microns. Capture of circulating melanoma cells may
provide the opportunity to study metastatic cells for basic understanding of the spread of cancer and to optimize patient