Kirk Shung

In vivo label-free photoacoustic microscopy of cell nuclei by excitation of DNA and RNA.

Imaging of cell nuclei plays a critical role in cancer diagnosis and prognosis. To image noninvasively cell nuclei in vivo without staining, we developed UV photoacoustic microscopy (UV-PAM), in which 266 nm wavelength UV light excites unlabeled DNA and RNA in cell nuclei to produce photoacoustic waves. We applied UV-PAM to ex vivo imaging of cell nuclei in a mouse lip and a mouse small intestine and to in vivo imaging of the cell nuclei in the mouse skin. The UV-PAM images of unstained cell nuclei match the optical micrographs of the histologically stained cell nuclei. Given intrinsic optical contrast and high spatial resolution, in vivo label-free UV-PAM has potential for unique biological and clinical application.

Integrated optical coherence tomography, ultrasound and photoacoustic imaging for ovarian tissue characterization

Ovarian cancer has the lowest survival rate of the gynecologic cancers because it is predominantly diagnosed in Stages III or IV due to the lack of reliable symptoms, as well as the lack of efficacious screening techniques. Detection before the malignancy spreads or at the early stage would greatly improve the survival and benefit patient health. In this report, we present an integrated optical coherence tomography (OCT), ultrasound (US) and photoacoustic imaging (PAI) prototype endoscopy system for ovarian tissue characterization. The overall diameter of the prototype endoscope is 5 mm which is suitable for insertion through a standard 5-12.5mm endoscopic laparoscopic port during minimally invasive surgery. It consists of a ball-lensed OCT sample arm probe, a multimode fiber having the output end polished at 45 degree angle so as to deliver the light perpendicularly for PAI, and a high frequency ultrasound transducer with 35MHz center frequency. System characterizations of OCT, US and PAI are presented. In addition, results obtained from ex vivo porcine and human ovarian tissues are presented. The optical absorption contrast provided by PAI, the high resolution subsurface morphology provided by OCT, and the deeper tissue structure imaged by US demonstrate the synergy of the combined endoscopy and the superior performance of this hybrid device over each modality alone in ovarian tissue characterization.

Impedance matching network for high frequency ultrasonic transducer for cellular applications

An approach for the design of an impedance matching network (IMN) for high frequency ultrasonic transducers with large apertures based on impedance analysis for cellular applications is presented in this paper. The main objectives were to maximize energy transmission from the excitation source to the ultrasonic transducers for cell manipulation and to achieve low input parameters for the safe operation of an ultrasonic transducer because the piezoelectric material in high frequency ultrasonic transducers is prone to breakage due to its being extremely thin. Two ultrasonic transducers, which were made of lithium niobate single crystal with the thickness of 15 μm, having apertures of 4.3 mm (fnumber=1.23) and 2.6mm (fnumber=0.75) were tested. L-type IMN was selected for high sensitivity and compact design of the ultrasonic transducers. The target center frequency was chosen as the frequency where the electrical admittance (|Y|) and phase angle (θz) from impedance analysis was maximal and zero, respectively. The reference center frequency and reference echo magnitude were selected as the center frequency and echo magnitude, measured by pulse-echo testing, of the ultrasonic transducer without IMN. Initial component values and topology of IMN were determined using the Smith chart, and pulse-echo testing was analyzed to verify the performance of the ultrasonic transducers with and without IMN. After several iterations between changing component values and topology of IMN, and pulse-echo measurement of the ultrasonic transducer with IMN, optimized component values and topology of IMN were chosen when the measured center frequency from pulse-echo testing was comparable to the target frequency, and the measured echo magnitude was at least 30% larger than the reference echo magnitude. Performance of an ultrasonic transducer with and without IMN was tested by observing a tangible dent on the surface of a plastic petridish and single cell response after an acoustic pulse was applied on a target cell.

Direct and sustained intracellular delivery of exogenous molecules using acoustic-transfection with high frequency ultrasound

Controlling cell functions for research and therapeutic purposes may open new strategies for the treatment of many diseases. An efficient and safe introduction of membrane impermeable molecules into target cells will provide versatile means to modulate cell fate. We introduce a new transfection technique that utilizes high frequency ultrasound without any contrast agents such as microbubbles, bringing a single-cell level targeting and size-dependent intracellular delivery of macromolecules. The transfection apparatus consists of an ultrasonic transducer with the center frequency of over 150 MHz and an epi-fluorescence microscope, entitled acoustic-transfection system. Acoustic pulses, emitted from an ultrasonic transducer, perturb the lipid bilayer of the cell membrane of a targeted single-cell to induce intracellular delivery of exogenous molecules. Simultaneous live cell imaging using HeLa cells to investigate the intracellular concentration of Ca2+ and propidium iodide (PI) and the delivery of 3 kDa dextran labeled with Alexa 488 were demonstrated. Cytosolic delivery of 3 kDa dextran induced via acoustic-transfection was manifested by diffused fluorescence throughout whole cells. Short-term (6 hr) cell viability test and long-term (40 hr) cell tracking confirmed that the proposed approach has low cell cytotoxicity.

Ultrasound Transducer Array Fabrication Based on Additive Manufacturing of Piezocomposites

Conventional methods for fabricating ultrasound imaging transducer arrays, especially for high frequency range (>20 MHz), are expensive, time consuming and limited to relatively simple geometries. In this paper, the development of an additive manufacturing (AM) process based on digital micromirror devices (DMDs) is presented for the fabrication of piezoelectric devices such as ultrasound transducer arrays. Both green-part fabrication and the sintering of fabricated greenparts have been studied. A novel two-channel design in the bottom-up projection system is presented to address the piezocomposite fabrication challenges including a small curing depth and viscous ceramic slurry recoating. A prototype system has been developed for the fabrication of green-parts with complex shapes and small features. Based on the fabricated green-parts, the challenges in the sintering process for achieving desired functionality are discussed. Various approaches for increasing the density of sintered components are presented. Dielectric and piezoelectric properties of the fabricated samples are measured and compared with those of bulk PZT samples. Based on the identified challenges in the DMD-based AM process, future work for achieving fully functional piezoelectric ceramic components is discussed.

Piezoelectric component fabrication using projection-based stereolithography of barium titanate ceramic suspensions

Purpose Conventional machining methods for fabricating piezoelectric components such as ultrasound transducer arrays are time-consuming and limited to relatively simple geometries. The purpose of this paper is to develop an additive manufacturing process based on the projection-based stereolithography process for the fabrication of functional piezoelectric devices including ultrasound transducers. Design/methodology/approach To overcome the challenges in fabricating viscous and low-photosensitive piezocomposite slurry, the authors developed a projection-based stereolithography process by integrating slurry tape-casting and a sliding motion design. Both green-part fabrication and post-processing processes were studied. A prototype system based on the new manufacturing process was developed for the fabrication of green-parts with complex shapes and small features. The challenges in the sintering process to achieve desired functionality were also discussed. Findings The presented additive manufacturing process can achieve relatively dense piezoelectric components (approximately 95 per cent). The related property testing results, including X-ray diffraction, scanning electron microscope, dielectric and ferroelectric properties as well as pulse-echo testing, show that the fabricated piezo-components have good potentials to be used in ultrasound transducers and other sensors/actuators. Originality/value A novel bottom-up projection system integrated with tape casting is presented to address the challenges in the piezo-composite fabrication, including small curing depth and viscous ceramic slurry recoating. Compared with other additive manufacturing processes, this method can achieve a thin recoating layer (as small as 10 μm) of piezo-composite slurry and can fabricate green parts using slurries with significantly higher solid loadings. After post processing, the fabricated piezoelectric components become dense and functional.

High-Frequency Ultrasonic Imaging with Lead-free (Na,K)(Nb,Ta)O3 Single Crystal:

Lead-free (Na,K)(Nb,Ta)O3 (KNNT) piezoelectric single crystal has been successfully grown using the top-seeded solution growth technique. The electromechanical coupling factors are very high (k33 = 0.827, kt = 0.646), and the dielectric loss tangent is as low as 0.004. Acoustic impedance was calculated to be 26.5 MRayl. From the single crystal, a single element transducer was fabricated. The transducer achieved a 57.6% −6 dB bandwidth and 32.3 µm axial resolution at the center frequency of 45.4 MHz, which can identify the cornea of porcine eyeball with high resolution. Comparison between KNNT single crystal and lead-based single crystal was discussed. The results suggest that this single crystal transducer is an excellent candidate to replace lead-containing transducer for high-frequency ultrasonic imaging applications.

Role of Excessive Autophagy Induced by Mechanical Overload in Vein Graft Neointima Formation: Prediction and Prevention

Little is known regarding the interplays between the mechanical and molecular bases for vein graft restenosis. We elucidated the stenosis initiation using a high-frequency ultrasonic (HFU) echogenicity platform and estimated the endothelium yield stress from von-Mises stress computation to predict the damage locations in living rats over time. The venous-arterial transition induced the molecular cascades for autophagy and apoptosis in venous endothelial cells (ECs) to cause neointimal hyperplasia, which correlated with the high echogenicity in HFU images and the large mechanical stress that exceeded the yield strength. The ex vivo perfusion of arterial laminar shear stress to isolated veins further confirmed the correlation. EC damage can be rescued by inhibiting autophagy formation using 3-methyladenine (3-MA). Pretreatment of veins with 3-MA prior to grafting reduced the pathological increases of echogenicity and neointima formation in rats. Therefore, this platform provides non-invasive temporal spatial measurement and prediction of restenosis after venous-arterial transition as well as monitoring the progression of the treatments.

Micromachined high frequency PMN-PT/Epoxy 1–3 composite ultrasonic annular arrays

Micromachined high-frequency PMN-PT 1–3 composite ultrasonic annular arrays have been developed. The design, fabrication, and performance of the miniature transducers are described in this paper. PMN-PT single crystal 1–3 composite was produced using micromachining techniques. The area of the single crystal pillar is 9 µm × 9 µm. The space between each pillar was ∼5 µm; this region was filled with a polymer. A six-element annular transducer with equal area for each element and 16 µm kerfs between annuli was produced. The composite thickness employed in these tests was 24 µm. A novel electric interconnection strategy for high density array elements is presented. After the transducer was attached to the electrical connection board and packaged, the array transducer was tested in a pulse/echo arrangement, whereby the center frequency, bandwidth, two-way insertion loss (IL), and cross talk between adjacent elements were measured for each annulus. The center frequency is about 50 MHz and the −6 dB bandwidth is about 90%. The minimum insertion loss is −19.5 dB at 50 MHz and the crosstalk between adjacent elements is about −35 dB. The micromachining techniques developed in this paper can be applied in fabrication of other array transducers.

Endoscopic ultrasound radial arrays fabricated with high-performance piezocrystal and piezocomposite

In this work, we develop the fabrication process of the radial array transducers, which are used to combine the gastroscope for acquiring ultrasound images of the surrounding organs, tissues and vessels. The PMNPT piezocrystal and its 1–3 piezocomposite were used as the active elements, respectively. High acoustic attenuation and flexibility composite material was used as the backing layers. The matching layers were specially designed according to the acoustic impedance of array elements. The pulse-echo responses acquired in water tank show that the PMNPT piezocrystal radial array transducer exhibits ∼80 % bandwidth and PMNPT/Epoxy 1–3 piezocomposite radial array transducer exhibits ∼100 % bandwidth, which has much broader bandwidth than commercial PZT array transducers (∼70 %). Thus, high resolution endoscopic ultrasonic images are expected for many clinical applications.