Novel contrasts in photoacoustic tomography

Photoacoustic tomography (PAT) combines rich optical contrast and high ultrasonic resolution in optically scattering tissue at depths. Taking advantage of its 100% sensitivity to optical absorption, PAT has been widely applied to structural, functional and molecular imaging, with both endogenous and exogenous contrasts, at superior depths than pure optical methods. This dissertation explores novel absorption contrast mechanisms of PAT based on optical/thermal patterns, endogenous cellular chromophores, nanoparticles, small-molecule dyes and genetically-encoded proteins. With these novel contrasts, the proof-of-concept applications of PAT have been extended to include homogenous flow measurements, targeted angiogenesis imaging and therapy, label-free white blood cell imaging, 3D-whole-organ cell nuclei imaging with a subcellular resolution, and in vivo neural activity imaging with voltage/calcium-sensitive indicators. Specifically, Chapter 1 introduces photoacoustic microscopy (PAM) and photoacoustic computed tomography (PACT) systems and discuss the motivation of the dissertation. Chapter 2 describes two photoacoustic (PA) flow measurement methods with optical and thermal patterns, which are applicable to homogenous flowing medium. In the first method, a Doppler frequency shift in PA signals of the flow was detected and used to calculate flow speeds. In the second method, unique features in an externally imposed thermal pattern of the flow, captured by repeated B-scans along the flow direction with a PAM system, revealed different flow speeds. Chapter 3 explores the unique PA contrast of macrophages, an important type of white blood cells. Macrophages were imaged by PAM without any label, and their measured PA spectrum was distinctive from the hemoglobin spectrum, so they can be potentially differentiated from red blood cells in the blood stream. Next, with a microtomy-assisted PAM system, cell nuclei distribution in whole organs, including mouse brain and mouse lung, were imaged with subcellular resolution. Chapter 4 introduces a type of target copper nanoparticles, which are less expensive and more biocompatible than its counterpart gold nanoparticles. The PA signals of neovasculature in the mouse flank were enhanced by the ___3-targeted copper nanoparticles. Moreover, the work shows the first example of a systemically targeted antiangiogenic drug delivery with a photoacoustic contrast nanoparticle in vivo. Chapter 5 demonstrates the voltage imaging capability of PA. A voltage sensitive dye with sufficient signal change was discovered and used as a PA voltage indicator for the first time. The mechanism was characterized through both PA imaging and spectroscopic methods. Its use was explored in a mouse epilepsy model and cortical electrical stimulation model in vivo. Finally, the deep imaging potential of PA was realized by imaging the voltage response of cells under 4.5 mm thick slice of rat brain tissue using a PACT system. Chapter 6 proves the neural calcium imaging capability of PA with a genetically encoded calcium indicator. In a fly model, I ambiguously demonstrated for the first time that PA can be used to imaging neural activities in the fly brain without the interference signals from hemoglobin. In the a live-mouse-brain-slice model, I successfully demonstrated the deep imaging capability of PA for calcium imaging by imaging through a 2-mm-thick scattering medium with a PACT system

Fast label-free multilayered histology-like imaging of human breast cancer by photoacoustic microscopy

The goal of breast-conserving surgery is to completely remove all of the cancer. Currently, no intraoperative tools can microscopically analyze the entire lumpectomy specimen, which results in 20 to 60% of patients undergoing second surgeries to achieve clear margins. To address this critical need, we have laid the foundation for the development of a device that could allow accurate intraoperative margin assessment. We demonstrate that by taking advantage of the intrinsic optical contrast of breast tissue, photoacoustic microscopy (PAM) can achieve multilayered histology-like imaging of the tissue surface. The high correlation of the PAM images to the conventional histologic images allows rapid computations of diagnostic features such as nuclear size and packing density, potentially identifying small clusters of cancer cells. Because PAM does not require tissue processing or staining, it can be performed promptly and intraoperatively, enabling immediate directed re-excision and reducing the number of second surgeries

Noninvasive photoacoustic microscopy of methemoglobin in vivo

Due to the various causes of methemoglobinemia and its potential to be confused with other diseases, in vivo measurements of methemoglobin have significant applications in the clinic. Using photoacoustic microscopy (PAM), we quantified the average and the distributed percentage of methemoglobin both in vitro and in vivo. Based on the absorption spectra of methemoglobin, oxyhemoglobin, and deoxyhemoglobin, three wavelengths were chosen to differentiate methemoglobin from the others. The methemoglobin concentrations calculated from the photoacoustic signals agreed well with the preset concentrations. Then we imaged the methemoglobin percentage in microtubes that mimicked blood vessels. Average percentages calculated for five samples with different methemoglobin concentrations also agreed well with the preset values. Finally, we demonstrated the ability of PAM to detect methemoglobin in vivo in a mouse ear. Our results show that PAM can quantitatively image methemoglobin distribution in vivo

Single-breath-hold photoacoustic computed tomography of the breast

We have developed a single-breath-hold photoacoustic computed tomography (SBH-PACT) system to reveal detailed angiographic structures in human breasts. SBH-PACT features a deep penetration depth (4 cm in vivo) with high spatial and temporal resolutions (255 µm in-plane resolution and a 10 Hz 2D frame rate). By scanning the entire breast within a single breath hold (~15 s), a volumetric image can be acquired and subsequently reconstructed utilizing 3D back-projection with negligible breathing-induced motion artifacts. SBH-PACT clearly reveals tumors by observing higher blood vessel densities associated with tumors at high spatial resolution, showing early promise for high sensitivity in radiographically dense breasts. In addition to blood vessel imaging, the high imaging speed enables dynamic studies, such as photoacoustic elastography, which identifies tumors by showing less compliance. We imaged breast cancer patients with breast sizes ranging from B cup to DD cup, and skin pigmentations ranging from light to dark. SBH-PACT identified all the tumors without resorting to ionizing radiation or exogenous contrast, posing no health risks

Structures of Sortase B from Staphylococcus aureus and Bacillus anthracis Reveal Catalytic Amino Acid Triad in the Active Site

Surface proteins attached by sortases to the cell wall envelope of bacterial pathogens play important roles during infection. Sorting and attachment of these proteins is directed by C-terminal signals. Sortase B of S. aureus recognizes a motif NPQTN, cleaves the polypeptide after the Thr residue, and attaches the protein to pentaglycine cross-bridges. Sortase B of B. anthracis is thought to recognize the NPKTG motif, and attaches surface proteins to m-diaminopimelic acid cross-bridges. We have determined crystal structure of sortase B from B. anthracis and S. aureus at 1.6 and 2.0 Å resolutions, respectively. These structures show a β-barrel fold with α-helical elements on its outside, a structure thus far exclusive to the sortase family. A putative active site located on the edge of the β-barrel is comprised of a Cys-His-Asp catalytic triad and presumably faces the bacterial cell surface. A putative binding site for the sorting signal is located nearby

Molecular Conformation Generation via Shifting Scores

Molecular conformation generation, a critical aspect of computational chemistry, involves producing the three-dimensional conformer geometry for a given molecule. Generating molecular conformation via diffusion requires learning to reverse a noising process. Diffusion on inter-atomic distances instead of conformation preserves SE(3)-equivalence and shows superior performance compared to alternative techniques, whereas related generative modelings are predominantly based upon heuristical assumptions. In response to this, we propose a novel molecular conformation generation approach driven by the observation that the disintegration of a molecule can be viewed as casting increasing force fields to its composing atoms, such that the distribution of the change of inter-atomic distance shifts from Gaussian to Maxwell-Boltzmann distribution. The corresponding generative modeling ensures a feasible inter-atomic distance geometry and exhibits time reversibility. Experimental results on molecular datasets demonstrate the advantages of the proposed shifting distribution compared to the state-of-the-art.Comment: 18 pages, 7 figure

Photoacoustic Doppler axial flow measurement of homogenous media using structured illumination

We propose time and frequency domain methods for homogenous flow measurement based on the photoacoustic Doppler effect. Excited by spatially modulated laser pulses, the flowing medium induces a Doppler frequency shift in the received photoacoustic signals. The frequency shift is proportional to the component of the flow speed projected onto the acoustic beam axis. These methods do not rely on particle heterogeneity in the medium. A red-ink phantom flowing in a tube immersed in water was used to validate the methods in both frequency and time domains