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PubMed2002

Scanning acoustic Doppler microscopy and scanning acoustic correlation microscopy.

Acoustic microscopy with vector contrast at 100 MHz in a fluid with immersed particles is used to detect the flow profile in front of a microscopic orifice. The velocity profile concerning the component in axial direction of the focused beam is derived from the phase contrast. Possibilities to resolve the flow profile also for the components in normal direction with respect to the axis are demonstrated. The methods concerning measurement techniques and data evaluation for scanning acoustic Doppler microscopy are presented. For scanning acoustic correlation microscopy the time dependent phase and amplitude signals resulting from sound waves scattered by the immersed particles (aluminium flakes with a typical diameter of 10 microm) have been analysed by correlation procedures. From the obtained autocorrelation functions the velocity distribution can be derived. Both methods can be applied simultaneously. Data analysis is based on the information contained in the originally obtained images in vector contrast derived from temporal and spatial resolved analogue and digital processing of the acoustic signals.

Journal Article↗
PubMed2002

A method of determining acoustical physical constants for piezoelectric materials by line-focus-beam acoustic microscopy.

We developed a new method of determining acoustical physical constants (elastic constant, piezoelectric constant, dielectric constant, and density) of piezoelectric materials with high accuracy. This method acquires velocities of leaky surface acoustic waves (LSAWs) excited on the water-loaded specimen surface, measured by line-focus-beam (LFB) acoustic microscopy, and bulk velocities of longitudinal and shear waves, measured with plane-wave transducers replacing the LFB device in the same system, together with the dielectric constants and density measured independently, for a small number of specimens. For LiNbO3 and LiTaO3 crystals, we demonstrated that we could accurately determine the constants by choosing proper propagation directions of LSAWs and bulk waves for three principal X-, Y-, and Z-cut specimens and one rotated Y-cut specimen [(104) plate for LiNbO3 and (012) plate for LiTaOa]. The accuracy is nearly the same as that for the constants determined only from the bulk wave velocities.

Journal Article↗
PubMed1979

Acoustic microscopy: resolution of subcellular detail.

Recent advances now permit the use of scanning acoustic microscopy for the analysis of subcellular components. By sequential viewing of identified fixed cells with acoustic, light, and electron microscopy, we have established that the acoustic microscope can readily detect such features as nuclei and nucleoli, mitochondria, and actin cables. Under optimal conditions, images can even be obtained of filopodia, slender projections of the cell surface that are approximately 0.1-0.2 micron in diameter. Small objects separated by as little as 0.5-0.7 micron can successfully be resolved. Three aspects of the acoustic micrographs prepared in this preliminary survey seem especially prominent. These are, first, the extraordinary level of acoustic contrast that can differentiate the various cytoplasmic organelles, even in regions of very thin cytoplasm; second, the reversals in acoustic contrast that occur when altering the plane of focus; and third, the sensitivity of the acoustic response to overall cytoplasmic thickness. The acoustic microscope uses a novel source of contrast that is based on local mechanical properties. In addition, it can provide a degree of resolution that is comparable to that of the light microscope.

Acoustics↗
PubMed1977

Acoustic microscopy of the human retina and pigment epithelium.

An acoustic microscope uses sound waves rather than light to image a sample, and displays viscoelastic rather than optical properties. The Stanford instrument, operating at frequencies near 1,000 MHz, achieves resolution and magnification that is comparable to a light microscope. Using this instrument, we examined sections of normal human retina and pigment epithelium and found that characteristic degrees of acoustic attenuation or phase shift were produced by structures such as cell nuclei, rod and cone outer segments, Bruch's membrane, red blood cells, and ocular pigment. Resolution was better with thin than thick sections, and fixation did not significantly alter the acoustic properties of the tissues studied. A comparison of iris tissue from albino and pigmented rabbits showed that melanin was a particularly strong acoustic attenuator. Acoustic microscopy may provide a new and direct means of probing the physical structure of tissues and cells.

Acoustics↗
PubMed1999

Tissue characterization of myocardial cells by use of high-frequency acoustic microscopy: differential myocyte sound speed and its transmural variation in normal, pressure-overload hypertrophic, and amyloid myocardium.

The purpose of the present study was to evaluate the acoustic properties of myocytes in normal, pressure-overload hypertrophic, and amyloid myocardium. Myocardial tissue specimens at autopsy were obtained from 10 subjects without cardiovascular disease, six patients with left ventricular (LV) hypertrophy, and six patients with cardiac amyloidosis. Sound speed of myocytes was measured at subendocardial and subepicardial regions in myocardium by use of a high-frequency (450 MHz) acoustic microscope. In normal myocardium, the sound speed of myocytes was significantly higher in subendocardial region (1,728+/-19 m/sec) than in subepicardial region (1,645+/-22 m/sec) (p<0.0001). A significantly higher sound speed of myocytes was observed in the subendocardial region in LV hypertrophic myocardium (1,779+/-19 m/sec) than that in normal myocardium (p<0.001). In amyloid myocardium, a significantly lower sound speed of myocytes was observed in subendocardial (1,560+/-8 m/sec) and subepicardial (1,594+/-48 m/sec) regions than that in respective regions of the normal myocardium (p<0.0001 and p<0.05, respectively). Transmural variation in sound speed of myocytes measured by high-frequency acoustic microscopy existed in normal left ventricle. The differential myocyte sound speed and its transmural variation was observed in LV hypertrophic and amyloid myocardium as compared with normal myocardium. High-frequency acoustic microscopy can be a promising technique for myocardial tissue characterization at the myocyte level.

Acoustics↗
PubMed2002

[Acoustic microscopy methods in studies of the filling materials microstructure].

The structure of filling materials (kemfil, polyacrylate cement, and Dentis, material produced by Stomadent Firm in Russia) at the interface with human dentin was studied in teeth subjected to retrograde filling with preliminary appendectomy. A prototype of table wide-field short pulse scanning acoustic microscope (50 mHz) (Acoustic Microscopy Center, Institute of Biochemical Physics of Russian Academy of Sciences) and Elsam acoustic microscope (200 mHz) (Leitz, Germany) were used. The results indicate that analysis of acoustic characteristics, including the data of acoustic microscopy, helps investigate the cement microstructure and evaluate the compactness and elasticity of samples; moreover, structural elements of the material, which are undetectable by other methods, are seen on acoustic images. These data can be used for evaluating the relationship between the microstructure and formation of mechanical properties, and, maybe, the patterns of cement interactions with dental tissues.

Acoustics↗
PubMed2000

Evaluation of progression in nonrheumatic aortic valvular stenosis by scanning acoustic microscopy.

To investigate distributions of hardness and thickness in nonrheumatic aortic stenosis (AS), scanning acoustic microscopy was used. The acoustic propagation speed (APS: m/s) and thickness at three sites (tip, middle and base) of aortic valve were measured in 18 cusps from 7 surgical patients with AS (late lesion), 27 showing mild lesions from 9 autopsy cases (early lesion) and 18 healthy from 6 autopsy cases (healthy). These were measured in each layer of cusps: fibrosa (F), spongiosa (S) or ventricularis (V). In early lesions, an increase in APS preceded the thickening and distributed in the tip (1666 +/- 107), the three layers of the middle (F: 1782 +/- 121; S: 1590 +/- 38; V: 1636 +/- 59) and the fibrosa of the base (1736 +/- 203). In late lesions, APS of the tip and three layers of the base were markedly increased. Progressive nonrheumatic AS is characterized by increased hardness that precedes the thickening, and its distribution may be related to mechanical stress.

Adult↗
PubMed1994

Subtraction scanning acoustic microscopy reveals motility domains in cells in vitro.

Scanning acoustic microscopy (SAM) observes all mechanical properties of living cells. Subtraction of the SAM images (SubSAM) of live cells was developed as a method for investigating minimal changes in cellular topography and elasticity. The image formation in the SubSAM takes into account the motion of cell mass as well as the changes of tension. High spatial and temporal resolution of the SubSAM revealed the structure of motile processes that develops at increasing time intervals, thus allowing the arising complexity of motion to be registered and investigated. Independent spots of activity emerge on a quiescent background as motility domains; they may change position, divide, merge, or disappear after a long time interval. In addition, zones of quiescence were identified over central parts of cytoplasmic lamellae. Nonmalignant (Ep: tadpole epidermal cells, XTH2: endothelial cells from tadpole hearts, 3T3 cells) and neoplastic cells (K2 cells of rat fibrosarcoma, A870N cells selected from K2) were investigated with the SubSAM. Three types of domains of subcellular cytoplasmic motility were identified in time series of two-dimensional SubSAM iamges in normal and neoplastic cells. Of them only the wave-like domain is self-evident, being derived from ruffling and protruding activity at the cell margin. Two other domains wait for detailed analysis. The oscillating domain is a visualization of tension within the cell(s), and the nucleating domain indicates intracellular processes possibly preceding locomotion. Differences in motile domains were found between low K2 and high A870N metastatic cells. The dynamics of motility domains of the A870N cells resembled that of the highly motile Ep cells. Cell morphotype and motile activity of the A870N cells are significantly influenced by the pH of the medium. It became evident that identification of the otherwise invisible motile domains in living cells by SubSAM opens a new approach to a characterization of cell motility in vitro and to an understanding of early cellular reactions to various stimuli.

Animals↗
PubMed2000

Synthetic aperture imaging by scanning acoustic microscopy with vector contrast

Based on phase sensitive scanning acoustic microscopy (PSAM), a novel scheme suitable for volume imaging has been developed. The method employs synthetic aperture insonification combined with synthetic aperture imaging. Excitation and detection are performed by planar scanning of two focusing transducer and vector (phase and amplitude) detectors for the ultrasonic wave packages observed in transmission. Examples for applications of the scheme, including technically relevant simplifications based on reduced dimensions of the scan ranges, are presented. Detection schemes already applied for non-destructive testing (NDT) and non-destructive evaluation (NDE) of the mechanical properties of functionally graded materials are an example for the application of the generalised approach presented here with reduced dimensions of the scan. The technique is suitable for NDT and NDE imaging respectively with three-dimensional resolution.

Journal Article↗
PubMed2001

Scanning acoustic microscopy study of human cortical and trabecular bone.

Scanning acoustic microscopy (SAM) has been used in the burst mode to study the properties of human cortical and trabecular bone. An Olympus UH3 SAM (Olympus Co., Tokyo, Japan) was used with a 400 MHz burst mode lens (120 degrees aperture, nominal lateral resolution 2.5 microm). The human cortical bone was from the midshaft of a femur from a 60+ male cadaver; the trabecular bone specimens were obtained from the distal femoral condyles of another 60+ human male cadaver. Elastic moduli for both trabecular and cortical bone were obtained by means of a series of calibration curves correlating SAM gray levels of known materials with their elastic moduli; specimens included: polypropylene, PMMA, Teflon, aluminum, Pyrex glass, titanium, and stainless steel. Values obtained by this method are in good agreement with those obtained by nanoindentation techniques. The three critical findings earlier by Katz and Meunier were observed here as well in both the cortical and trabecular bone samples.

Artifacts↗
PubMed1988

Acoustic microscopy of red blood cells.

We used a scanning acoustic microscope to image normal and outdated red blood cells, cells with different hemoglobin content, red cell ghosts, and cells treated with various drugs that induce echinocyte-stomatocyte transformation. Images were obtained at different planes of focus within the cells, corresponding to maxima and minima of signal intensity. Digitization and gray scale amplitude mapping were used to create axonometric plots that display signal amplitude variations within the cells. The images of red cells contain features produced by differences in topology, density, elasticity, and absorption. Both hemoglobin content and the cell cytoskeleton contribute to image features, and various deformations, characterized by the formation of blebs and vacuoles, are displayed in cells undergoing echinocyte-stomatocyte transformation. These preliminary findings, although mainly descriptive, indicate that acoustic microscopy may be a useful new method for evaluating red cell deformation and associated changes in mechanical properties.

Erythrocyte Aging↗
PubMed2006

Does decalcification alter the tissue sound speed of rabbit supraspinatus tendon insertion? In vitro measurement using scanning acoustic microscopy.

Failure of the tendon or ligament insertions is one of the most common injuries in the Orthopaedic field. To elucidate the pathogenesis of those injuries, the authors had attempted to measure the tissue sound speed that could be correlated to its elasticity using scanning acoustic microscopy (SAM). For the application of SAM to tendon or ligament insertions, it was necessary to determine the role of decalcification in SAM measurements since mineralized tissues including bone or mineralized fibrocartilage were present at the insertion site. To assess whether decalcification alters the tissue sound speed or not, supraspinatus tendon insertion of six Japanese white rabbits were measured with SAM operating in the frequency range of 50-150 MHz. Right supraspinatus tendons attached to the humeral head were cut into two pieces at the center of the tendon. Then, they were fixed with 10% neutralized formalin for 12 h. In each specimen, medial half was not decalcified, while lateral half was decalcified with ethylene-diamine-tetra-acetic acid (EDTA). After embedding in paraffin, 5 microm thick specimens were prepared for the measurement using SAM. The mean sound speed in each histologic zone was evaluated, and subsequently compared to that measured in the undecalcified and the decalcified specimens. Mean sound speed of non-mineralized fibrocartilage was 1544 m/s in the undecalcified specimens, while the value of 1541 m/s was determined in the decalcified ones. On the other hand, it decreased 2-3% after decalcification in the mineralized tissue including mineralized fibrocartilage and bone (mineralized fibrocartilage: undecalcified = 1648 m/s, decalcified = 1604 m/s; bone: undecalcified = 1716 m/s, decalcified = 1677 m/s). However, no significant differences were found between the undecalcified and the decalcified specimens (non-mineralized fibrocartilage: p = 0.84, mineralized fibrocartilage: p = 0.35, bone: p = 0.28). These results indicate that SAM could be applied to determine the properties of the tendon or the ligament insertions after decalcification with EDTA. Although SAM is applicable only for in vitro experimental study, it is expected that these data will contribute to better understanding concerning the biomechanics of tendon or ligament insertions as well as the pathogenesis of their failure at a microscopic level.

Animals↗
PubMed2000

Comparison of left ventricular diastolic filling with myocyte bulk modulus using Doppler echocardiography and acoustic microscopy in pressure-overload left ventricular hypertrophy and cardiac amyloidosis.

BACKGROUND: The myocardial bulk modulus has been described as the constitutive properties of the left ventricular (LV) wall and is measured as rho V2 (rho = density, V = sound speed) using acoustic microscopy. HYPOTHESIS: The study was undertaken to assess the relationship between the myocyte bulk modulus and transmitral inflow patterns in patients with pressure-overload LV hypertrophy (LVH) and cardiac amyloidosis (AMD). METHODS: In 8 patients with LVH, 8 with AMD, and 10 controls without heart disease, the transmitral inflow pattern was recorded by Doppler echocardiography before death, and myocardial tissue specimens were obtained at autopsy. The tissue density and sound speed in the myocytes were measured by microgravimetry and acoustic microscopy, respectively. The diameters of the myocytes were measured on histopathologic specimens stained by the elastica Van Gieson method. RESULTS: In the subendocardium, the myocyte bulk modulus was larger in LVH (2.98 x 10(9) N/m2, p < 0.001) and smaller in AMD (2.61 x 10(9) N/m2, p < 0.001) than in the controls (2.87 x 10(9) N/m2). The myocyte diameter in LVH (26 +/- 1 microns) was larger than that in the control (21 +/- 1 microns, p < 0.001) and AMD (20 +/- 1 microns, p < 0.001). The bulk modulus in the subendocardial myocyte significantly correlated with the deceleration time (DT) of the early transmitral inflow (r = 0.689, p = 0.028 in control, r = 0.774, p = 0.024 in LVH, and r = 0.786, p = 0.021 in AMD). CONCLUSION: The changes in the myocyte elasticity as represented by the bulk modulus were limited to the subendocardial layers and may be related to relaxation abnormalities in LVH and a reduction in LV compliance in AMD.

Acoustics↗
PubMed1984

Acoustic microscopy--principles and applications in the studies of biomaterial microstructure.

The acoustic microscopy is based on totally different physical concepts than both the optical and electron microscopes. Therefore, the information obtained with an acoustic microscope is significantly different from those obtainable by the other two. By employing elastic waves, detailed microscopic information regarding biomaterial properties like density, elasticity, viscosity and viscoelasticity are obtained with an acoustic microscope. Live tissue can be examined without fixing or chemical staining. The Scanning Laser Acoustic Microscope (SLAM) and the Scanning Acoustic Microscope (SAM) are the only acoustic microscopes commercially available today. The SAM operates at as high as 4.2 GHz frequency providing at least five times better resolution than the optical limit. The SLAM, however, operates generally at 100 or 500 MHz and can therefore examine thicker samples than the SAM, but at lower resolutions. For optically translucent samples, the SLAM can also provide an optical image simultaneously with the acoustical one. A laser beam is employed in the scanning mechanism in the SLAM, compared to mechanically translating the sample in the SAM. Consequently, the scanning rate is much faster in the SLAM at about 30 frames per second, compared to several seconds per frame for the SAM. Both qualitative as well as quantitative information is obtainable about the material under examination. As a qualitative tool, the information is used to classify and sort materials and detect and localize flaws and defects in optically opaque samples. A microscopic map of the specimen's mechanical properties can be produced using information from the acoustic image. This information may then lead to better understanding of biomaterial microstructure and also can be a valuable aid in characterization of certain subtle morphological differences.

Animals↗
PubMed1981

Acoustic microscopy of living cells.

This paper reports preliminary results of the observation by acoustic microscopy of living cells in vitro. The scanning acoustic microscope uses high-frequency sound waves to produce images with submicrometer resolution. The contrast observed in acoustic micrographs of living cells depends on the acoustic properties (i.e., density, stiffness, and attenuation) and on the topographic contour of the cell. Variation in distance separating the acoustic lens and the viewed cell also has a profound effect on the image. When the substratum is located at the focal plane, thick regions of the cell show a darkening that can be related to cellular acoustic attenuation (a function of cytoplasmic viscosity). When the top of the cell is placed near the focal plane, concentric bright and dark rings appear in the image. The location of the rings can be related to cell topography, and the ring contrast can be correlated to the stiffness and density of the cell. In addition, the character of the images of single cells varies dramatically when the substratum upon which they are grown is changed to a different material. By careful selection of the substratum, the information content of the acoustic images can be increased. Our analysis of acoustic images of actively motile cells indicates that leading lamella are less dense or stiff than the quiescent trailing processes of the cells.

Animals↗
PubMed2005

Characterization of polymer thin films by phase-sensitive acoustic microscopy and atomic force microscopy: a comparative review.

The potential of phase-sensitive acoustic microscopy (PSAM) for characterizing polymer thin films is reviewed in comparison to atomic force microscopy (AFM). This comparison is based on results from three-dimensional vector contrast imaging and multimodal imaging using PSAM and AFM, respectively. The similarities and differences between the information that can be derived from the AFM topography and phase images, and the PSAM phase and amplitude micrographs are examined. In particular, the significance of the PSAM phase information for qualitative and quantitative characterization of the polymer films is examined for systems that generate surface waves, and those that do not. The relative merits, limitations and outlook of both techniques, individually, and as a complementary pair, are discussed.

Journal Article↗