Dual-frequency Ultrasound Research Articles

Microbubble detection following hyperbaric chamber dives using dual-frequency ultrasound

Venous gas emboli (VGE) can be readily detected in the bloodstream using existing ultrasound methods. No method currently exists to detect decompression-induced microbubbles in tissue. We hypothesized that dual-frequency ultrasound (DFU) could detect these microbubbles. With DFU, microbubbles are driven with two frequencies: a lower "pump" (set to the resonant frequency of the desired bubble size) and a higher "image" frequency. A bubble of the resonant size emits the sum and difference of the two transmitted frequencies. For this study we used a pump frequency of 2.25 MHz and an image frequency of 5.0 MHz, which detects bubbles of roughly 1-10 μm in diameter in a water tank. Four anesthetized swine were pressurized at 4.5 ATA for 2 h and decompressed over 5 min, inducing moderate to very severe VGE scores. Four sites on the thigh of each swine were monitored with DFU before and after the dives. A single mock dive was also performed. The number of sites returning signals consistent with microbubbles increased dramatically after the chamber dive (P < 0.01), but did not change with the mock dive. The increase in DFU signal after the chamber dive was sustained and present at multiple sites in multiple swine. This research shows for the first time that decompression-induced tissue microbubbles can be detected using DFU and that DFU could be used to monitor decompression-induced microbubbles at multiple sites on the body. Additionally, DFU could be used to track the time course of microbubble formation and growth during decompression stress.

Effects of non-optimal focusing on dual-frequency ultrasound measurements of bone

In pulse-echo (PE) ultrasound measurements, the use of focused transducers is desirable for quantitative assessment of bone characteristics because of the attenuation in the overlying soft tissues. However, the variable thickness and composition of the soft tissue overlying bone affect the focal depth of the ultrasound beam and induce errors into the measurements. To compensate for the attenuation-related effects caused by the interfering soft tissue (i.e., fat and lean tissue), a dual-frequency ultrasound (DFUS) technique was recently introduced. The aim of this study was to investigate the effect of non-optimal focal depth of the ultrasound beam on the determination of the integrated reflection coefficient (IRC) of bone when overlaid by an interfering layer composed of oil and water. The feasibility of the DFUS-based correction of the IRC was evaluated through numerical simulations and experimental measurements. Even when the interfering layer-bone interface was out of focus, the total thickness of the interfering layer could be accurately determined with the technique. However, based on the simulations, the errors in the determination of the composition of the interfering layer increased (0.004 to 113.8%) with the increase in distance between the interfering layer-bone interface and the focus of the ultrasound beam. Attenuation compensation, based on the true composition of the interfering layer, resulted in an average relative error of 22.3% in the IRC values calculated from the simulations. Interestingly, the attenuation compensation with the interfering layer composition estimated using the DFUS technique resulted in a smaller average relative error of 14.9% in the IRC values. The simulations suggest that DFUS can reduce the errors induced by soft tissue in bone PE ultrasound measurements. The experimental measurements indicate that the accuracy of the IRC measurements is rather similar when using DFUS correction or correction based on the true composition of the interfering layer. However, the results suggest that accurate determination of soft tissue composition may be difficult without optimal focusing of the ultrasound beam on the soft tissue-bone interface.

Bone diagnostics using dual frequency ultrasound measurements.

Diagnosis of osteoporosis is made at skeletal sites composed mainly of trabecular bone. Calcaneus has been the first location for through transmission (TT) ultrasonic measurements of trabecular bone. Similarly as with the DXA, the best prediction of the hip fractures could be obtained by making the measurements at hip. To realize axial pulse-echo (PE) ultrasound measurements, e.g., in hip we have introduced the dual frequency ultrasound (DFUS) technique to minimize the effects of soft tissues overlying the bone and to correct the PE-parameters [Riekkinen et al., Ultrasound Med. Biol. 34, 1703–1708 (2008)]. Based on our experimental measurements at frequencies of 2.25 and 5.0 MHz and finite difference simulations, the DFUS technique detects minor changes in bone density, despite variable composition of soft tissue. For TT-geometry, the composition of the interfering soft tissues may also be solved with the DFUS, e.g., by measuring the reflection from the bone surfaces at both sides of the bone. The measurements with phantom materials indicated that accuracy of TT-parameters, such as broadband ultrasound attenuation, were significantly improved by DFUS. Potentially, the DFUS could also benefit the in vivo TT-measurements of trabecular bone. [Support from Academy of Finland is acknowledged.]

Numerical Analysis of Uncertainties in Dual Frequency Bone Ultrasound Technique

Quantitative ultrasound (QUS) measurements are used in the diagnostics of osteoporosis. However, the variation in the thickness and composition of the overlying soft tissue causes significant errors to the bone QUS parameters and diminishes the reliability of the technique in vivo. Recently, the dual frequency ultrasound (DFUS) technique was introduced to minimize the errors related to soft tissue effects. In this study, the significance of soft tissue induced errors and their elimination with the DFUS technique were simulated using the finite difference time domain technique. Furthermore, we investigated the potential of the DFUS corrected integrated reflection coefficient (IRC) of bone to detect changes in the cortical bone density. The effects of alterations in the thickness of fat and lean tissue layers and the inclination between the soft-tissues and between the soft tissue-bone layers were simulated. When the angle of the soft tissue interface was zero, i.e., perpendicular to the incident ultrasound beam, the DFUS-calculated soft tissue composition correlated highly linearly with the true soft tissue composition. The inclination between the soft tissue-bone layers was found to be critical. Even a 2-degree inclination between the soft tissue and the bone surface induced an almost 18% relative error in the corrected IRC. Increasing the inclination between the soft tissue layers increased the error in the DFUS-calculated lean and fat tissue thickness. This error was especially significant at inclination angles greater than 20 degrees. The significant soft tissue induced errors in IRC values (>300 %) could be effectively minimized (<10 %) by means of the DFUS correction. Importantly, after the DFUS correction, physiologically relevant variation in the cortical bone density could be detected ( p < 0.05). (E-mail: Markus.Malo@uef.fi)

Signals consistent with microbubbles detected in legs of normal human subjects after exercise

Exercise may produce micronuclei (presumably gas-filled bubbles) in tissue, which could serve as nucleation sites for bubbles during subsequent decompression stress. These micronuclei have never been directly detected in humans. Dual-frequency ultrasound (DFU) is a resonance-based, ultrasound technique capable of detecting and sizing small stationary bubbles. We surveyed for bubbles in the legs of six normal human subjects (ages 28-52 yr) after exercise using DFU. Eleven marked sites on the left thigh and calf were imaged using standard imaging ultrasound. Subjects then rested in a reclining chair for 2 h before exercise. For the hour before exercise, a series of baseline measurements was taken at each site using DFU. At least six baseline measurements were taken at each site. Subjects exercised at 80% of their age-adjusted maximal heart rate for 30 min on an upright bicycle ergometer. After exercise, the subjects returned to the chair, and multiple postexercise measurements were taken at the marked sites. Measurements continued until no further signals consistent with bubbles were returned or 1 h had elapsed. All subjects showed signals consistent with bubbles after exercise at at least one site. The percentage of sites in a given subject showing signals significantly greater than baseline (P < 0.01) at first measurement ranged from 9.1 to 100%. Overall, 58% of sites showed signals consistent with bubbles at the first postexercise measurement. Signals decreased over time after exercise. These data strongly suggest that exercise produces bubbles detectable using DFU.

Treatment of Murine Tumor Models of Breast Adenocarcinoma by Continuous Dual-Frequency Ultrasound

Introduction: Acoustic transient cavitation is the primary mechanism of sonochemical reaction and has potential use for tumor treatment. In this study, the in vivo anti-tumor effect of simultaneous dual-frequency ultrasound at low-level intensity (ISATA < 6 W/cm2) was investigated in a spontaneous murine model of breast adenocarcinoma in Balb/c mice. Materials and Methods: Forty tumor bearing mice were divided into four groups (10 in each group). The treated groups received 15 or 30 minutes of combined dual-frequency ultrasound in continuous mode (1 MHzcon + 150 kHzcon) respectively. The control and the sham groups contained the untreated mice. The tumor growth delay parameters including tumor volume, relative tumor volume, T5 and T2 (the needed time for each tumor to reach 5 and 2 times the initial tumor volume, respectively), survival period and percent of tumor growth inhibition ratio were measured on different days after treatment. Results: The results showed that the 30 min treatment was effective in tumor growth delay and percent of tumor growth inhibitory ratio compared to the sham and the control groups. The tumor volume growth and relative volume of tumors in the same treated group showed an anti-tumor effect relative to the sham and the control groups. There was a significant difference in tumor volume growth between this 30 min treatment group and the sham group 12 days after treatment (p-value <0.05). The mean of the survival period for animals in the 30 min treatment group was 16% more than the control group. The percent of tumor growth inhibition in the 30 and 15 min treatment groups were 23% and 20% respectively, showing a statistically significant difference. Conclusion: Sonodynamic therapy with combined dual–frequency ultrasound in progressive wave mode can be useful in cancer therapy.

Treatment of Murine Tumors Using Dual-Frequency Ultrasound in an Experimental in Vivo Model

Acoustic inertia cavitation is the primary mechanism underlying sonochemical reactions and has potential for use in tumor treatment. In in vitro experiments that were performed previously and are thus not included in this paper, we found that the ultrasonically-induced chemical reactions are greatly accelerated when ultrasound is simultaneously applied at frequencies of 1 MHz and 150 kHz.. In this study, the in vivo anti-tumor effect of the simultaneous dual-frequency ultrasound at low level intensity (I SPTA <6 W/cm 2) was investigated in a murine model of breast adenocarcinoma in Balb/c mice. The tumor-bearing mice were divided into five groups: those treated with combined dual-frequency ultrasound in continuous mode (1 MHz con+150 kH zcon) for 30 and 15 min (C and D), those treated with dual-frequency ultrasound in which the source of 1 MHz was in pulse mode (duty cycle of 80%) and that of 150 kHz was in continuous mode for 30 min (E), and untreated control and sham groups (A and B). The tumor growth parameters evaluated to assess delay include tumor volume, relative tumor volume, and T 5 and T 2, which are the time needed for each tumor to reach 5 and 2 times its initial volume, respectively. The survival period and percent of tumor growth inhibition ratio and were measured at various times after treatment. The results show that treatment with a combined continuous mode of 1 MHz con+150 kHz con and a pulse mode of 1 MHz pl,80%+150 kHz con effectively delayed tumor growth and increased the tumor growth inhibitory ratio compared to the sham group. When the tumor volume growth and relative volume of tumors in treated groups C, D and E were examined, an anti-tumor effect was observed in groups E and C. There is a significant difference between groups E and C and the sham group 12 d after treatment for tumor volume growth and 18 d after treatment for relative tumor volume ( p < 0.05). The mean survival periods for animals in groups C and E were 16% and 17% more than the control group. T 5 and T 2 (in days) of groups E, C and B are 19.1 ± 0.4 and 7.6 ± 1.5, 17.2 ± 0.6 and 6.3 ± 2.1, 14.3 ± 1.1 and 5.1 ± 2.5 d, respectively. There was a significant difference between groups C and E and the sham group (B) for T 5 ( p < 0.05), but for T 2, there was no significant difference between groups ( p > 0.05). The tumor growth inhibition ratio in groups C, D and E was 23, 20 and 37% of that in group B, and those differences are statistically significant ( p <0.05). It is concluded that sonodynamic therapy with combined dual–frequency ultrasound in a progressive wave mode can be useful for cancer therapy.

Dual-frequency ultrasound technique minimizes errors induced by soft tissue in ultrasound bone densitometry

Most bone ultrasound devices are designed for through-transmission measurements of the calcaneus. In principle, ultrasound backscattering measurements are possible at more typical fracture sites of the central skeleton. Unfortunately, soft tissue overlying the bones diminishes reliability of these measurements. To apply the single-transducer dual-frequency ultrasound (DFUS) technique to eliminate the errors induced by soft tissue on the measurements of integrated reflection coefficient (IRC) in human distal femur in vivo. Ultrasound and dual-energy X-ray absorptiometry (DXA) examinations were conducted on a bodybuilder during a 21-week training and dieting period. Significant changes in quantity and composition of soft tissue took place during the diet. However, DXA measurements showed no significant effects on bone density measurements. The single transducer DFUS technique enabled the determination of local soft-tissue composition, as verified by comparison with the DXA (r=0.91, n=8, p<0.01). Further, the technique eliminated the soft-tissue-induced error from IRC measured for the bone. The uncorrected IRC associated significantly with the change in local soft-tissue composition (r=-0.83, n=8, p<0.05), whereas the corrected IRC values showed no significant dependence (r=-0.30, n=8, p=0.46) on local soft-tissue composition. The DFUS technique may significantly enhance the accuracy of clinical ultrasound measurements of bone.

Sonochemistry and Sonoluminescence under Dual-Frequency Ultrasound Irradiation in the Presence of Water-Soluble Solutes

The sonoluminescence emission and sonochemical efficiency of a typical laboratory scale, high-frequency standing wave reactor (355 kHz), stimulated by low-frequency pulses (20 kHz) has been investigated in the presence and absence of some water-soluble solutes, namely, propanol and polyethylene oxide. It has been found that, although dual-frequency sonication causes a decrease in the integrated sonoluminescence intensity and sonochemical efficiency in water, in the presence of solutes, a significant enhancement in activity could be attained. This enhancement effect is ascribed, in part, to changes in the extent of bubble coalescence brought about by the water-soluble solutes.

Dual-Frequency Ultrasound—New Pulse–Echo Technique for Bone Densitometry

Quantitative ultrasound has been suggested for screening of osteoporosis. Most commercial ultrasound devices are based on the through-transmission measurement of calcaneus, which is not a typical fracture site. In contrast to through-transmission measurements, reflection and backscattering measurements may be conducted at typical fracture sites such as vertebra and proximal femur. At these regions, soft tissues overlying bones affect reliability of the measurements. In this study, a novel dual-frequency ultrasound (DFUS) pulse–echo technique is introduced for reduction of the errors induced by soft tissues. First, DFUS was validated using elastomer samples. For further validation, human trabecular bone samples ( n = 25) covered with heterogeneous soft tissues were measured at frequencies of 2.25 MHz and 5.0 MHz. The DFUS technique reduced ( p < 0.01) the mean error induced by soft tissue from 58.6% to –4.9% and from 127.4% to 23.8% in broadband ultrasound backscattering and integrated reflection coefficient (at 5.0 MHz), respectively. To conclude, the DFUS, being the first ultrasound technique capable of determination of the composition and thickness of the soft tissue overlying the bone, may enhance the accuracy of clinical ultrasound measurements. Thereby, DFUS shows a significant clinical potential. (E-mail: Ossi.Riekkinen@uku.fi)

Dual frequency ultrasound measurement of bone ‐ a technique for elimination of soft tissue effects on pulse‐echo measurements

Quantitative ultrasound (US) measurements have been suggested for screening of osteoporosis. However, soft tissues overlying bones affect reliability of the measurements. In this in vitro study, a novel dual frequency ultrasound (DFUS) technique is introduced for elimination of the errors induced by soft tissues on bone pulse‐echo US measurements. In DFUS, US reflection from soft tissue‐bone interface is measured with two different US frequencies. By knowing the frequency specific US attenuation and speed in adipose and lean tissues, the effect of soft tissue can be determined. DFUS, conducted at frequencies of 2.25 MHz and 5.0 MHz, was validated using human trabecular bone samples (n = 25) covered with heterogeneous soft tissues. DFUS, reduced (p < 0.01) the mean error induced by soft tissues from 58.6% to ‐4.9% and from 127.4% to 23.8% in broadband ultrasound backscattering and integrated reflection coefficients (at 5.0 MHz), respectively. Our results suggest that DFUS is a technique capable to minimize...

Dual frequency ultrasound‐ A pulse‐echo technique for analysis of layered material

Ultrasound (US) thickness gauges typically analyse layered materials by utilizing ultrasound reflections between different layers and prior knowledge for the material order within the layered structure. In this study, a dual frequency ultrasound (DFUS) technique is applied to eliminate the effect of overlying layered structure on the measurements of the object of interest without prior knowledge of the order of materials within the multilayered structure. DFUS technique utilizes prior knowledge on US attenuation coefficient and speed at two frequencies in multilayered materials, consisting of two different material types. Then, US reflection from the front (first) and the back (last) surfaces of the multilayered structure is measured using two different US frequencies. No reflections from the internal interfaces are needed. The technique was validated using several elastomer samples and their combinations, measured at 2.25 MHz and 5.0 MHz. DFUS reduced the mean error, induced by the overlying elastomers, in reflection from the object of interest from 103.6 ‐ 289.4% to ‐15.9 ‐ 5.6% and from 37.5 ‐ 77.5% to ‐12.0 ‐ 4.9% with 5.0 MHz and 2.25 MHz, respectively. Based on these results, DFUS is a straightforward technique to analyse the multilayered structure without the need for echoes from internal interfaces.

Dual frequency ultrasound technique enables determination of soft tissue composition and improves reliability of in vivo ultrasound bone densitometry

Soft tissues diminish reliability of the bone ultrasound backscatter measurements. In this study, the ability of a single broadband transducer dual frequency ultrasound (DFUS) technique to monitor the changes in soft tissue was investigated in a body builder during a 21 week training and dieting period, inducing a weight loss of 16.5 kg (18%). Then, DFUS was applied to correct the errors induced by soft tissues on the measurements of integrated reflection coefficient (IRC) in human distal femur. In DFUS, US reflection from soft tissue‐bone interface is determined with two different US frequencies and, by knowing the frequency specific US attenuation and speed in adipose and lean tissues, their content can be determined. The dual energy X‐ray absorptiometry (DXA) indicated that significant changes in quantity and composition of soft tissue, but not in bone density, took place during the diet. As compared with DXA, the single transducer DFUS could determine local soft tissue composition (r2 = 0.88, n=8, p <...