Stable Microbubbles Research Articles

Incorporating ultrasound contrast in the laboratory: A series on contrast echocardiography, article 1

Contrast echocardiography, in the form of agitated saline solution, has been routinely performed for more than 30 years. Because these large and fragile microbubbles do not pass through the pulmonary circulation, they are visible only on the right side of the heart and are typically used to rule out an atrial level shunt. Recent advances in second-generation agents have produced smaller and more stable microbubbles that can pass through the pulmonary circulation intact and that now can provide left ventricular (LV) opacification with a peripheral intravenous injection. As the number of Food and Drug Administration–approved ultrasonographic (ultrasound) contrast agents increases, contrast echocardiography will continue to move at an increasing pace from research laboratories to routine clinical use. There is more to the successful implementation of contrast than simply ordering a few vials of an agent. New challenges exist for the cardiac sonographer, as well as other team members, as ultrasound contrast echocardiography moves into routine clinical use. In this first in a series of articles, basic information regarding what laboratories need to integrate contrast echocardiography will be discussed in detail.

Aplicaciones del contraste en la ecocardiografía

The development of contrast media in ultrasound has been slow and sporadic, and there are no fully satisfactory agents for clinical imaging to date. Most contrast agents consist of air filled microbubbles which generate scattered echoes and enhance the ultrasound information. In this article we review the different phases of contrast echocardiography their potentials and clinical applications. First, right-sided echocontrast which is mainly used for the assessment of intracardiac shunts. Second, left-sided contrast agents with smaller and more stable microbubbles, that allow the visualization of the left ventricle after intravenous injection. With these agents, one it is now possible to study myocardial perfusion which is one of the most attractive potentials of these types of agents.

The Stable Microbubble Test on Tracheal Aspirate Samples from Newborn Babies for Diagnosis of Surfactant Deficiency and/or Surfactant Malfunction

For predicting the risk of respiratory distress syndrome (RDS), lung surfactant in amniotic fluid can be assessed by observation of stable microbubbles (<15 µm diameter). It was investigated if the stable microbubble test (MT) developed on amniotic fluid could be applied on tracheal aspirate samples (TA) obtained from newborn infants. Forty-four TA from 29 newborn infants without pulmonary diseases (group 1) and 65 TA from 21 newborn infants with respiratory insufficiency (group 2) were analyzed by the MT for predicting surfactant deficiency and/or surfactant malfunction. Ten microbubbles/mm² was defined as cutoff value of the MT. A specificity of 90% and a sensitivity of 52% was obtained with this cutoff value. A much lower rate of false negative results (sensitivity 91%) was noted after surface tension measurement of TA. The sensitivity of the MT (52%) does not encourage the use of the MT on TA.

New developments in ultrasound systems for contrast echocardiography.

Contrast echocardiography (CE) has evolved significantly in the past decade. Contrast agents and the hardware and software used to detect them and display optimal images have developed in tandem. Not only are hardware and contrast agents available that allow left ventricular cavity enhancement, but recent research points to the usefulness of CE for the evaluation of myocardial perfusion in the cardiac catheterization laboratory and operating room. Advances in ultrasound technology, such as transient harmonic imaging and integrated backscatter, coupled with the development of newer contrast agents that contain smaller, more stable microbubbles capable of transpulmonary passage for intravenous injection, promise a vast increase in the applications of CE in clinical practice.

Influence of dissolved gas on optical breakdown and small-angle scattering of light in liquids

It is demonstrated that the small-angle laser beam scattering pattern in liquid depends greatly on the dissolved gas content: the intensity of the scattering steeply drops after degassing doubly distilled water. It was experimentally found that the parameters or small-angle scattering of light correlate strongly with those of optical breakdown in liquid. Namely, the fractal dimension and the gyration radius of scatterers in aqueous KCl solutions behave similar to the optical breakdown probability in those liquids versus salt concentration, while the optical breakdown cannot be induced after degassing. The effect might be associated with the presence of stable gas microbubbles in liquid.

Surfactant proteins and stable microbubbles in tracheal aspirates of infants with respiratory distress syndrome: relation to the degree of respiratory failure and response to exogenous surfactant.

Surfactant proteins (SP-A and SP-BC), albumin (ALB), and stable microbubble (SM) count were measured in tracheal aspirates from infants with respiratory distress syndrome (RDS) receiving single-dose Surfactant-TA (surfactant group, n = 32) or no surfactant (control group, n = 12), and those without RDS (non-RDS group, n = 8) to determine biochemical and biophysical status of surfactant in the course of RDS after surfactant replacement. Surfactant therapy resulted in immediate and sustained elevations of SP-BC/ALB and SM count with a rapid fall in ventilatory index to levels measured in the non-RDS group, whereas these indices improved slowly in the control group. The SP-A/ ALB was initially low in both RDS groups and increased to levels measured in the non-RDS group by age 48 h. Multiple regression analysis showed that SP-BC/ALB, postnatal age, SM count, SM count/SP-A plus SP-BC, and surfactant therapy were independently associated with the severity of RDS as assessed by ventilatory index (r = 0.75, P < 0.0001; number of samples = 256). Infants with a relapse response to surfactant (n = 9) had levels of SP-A/ALB and SP-BC/ALB similar to those measured in the sustained group (n = 23), but had significantly lower SM count and SM count/SP-A plus SP-BC between 24 and 96 h of age. Surfactant therapy normalizes the surfactant and respiratory status of infants with RDS. Surfactant dysfunction rather than depletion may explain the relapse response seen in some surfactant recipients.

Ethanol resistive microbubble test: a modification of the stable microbubble test used to predict respiratory distress syndrome.

The stable microbubble (SM) test on gastric aspirate obtained at birth proved useful in identifying infants who would develop respiratory distress syndrome (RDS). This test involves only the count of stable microbubbles of < or = 15 microns in diameter. Larger bubbles (> 15 microns in diameter) are not necessary for the test and may interfere with stable microbubble counting. The aims of the present study were to determine: (i) if larger bubbles could be selectively removed by adding ethanol, a potent bubble breaker; and (ii) if the predictive value of this modified test, the ethanol resistive microbubble (ERM) test, on the development of RDS was similar to that of the SM test. Varying amounts of different concentrations of ethanol-water solutions were added to the top of the bubble crop generated by the SM test procedure, and the mean counts of stable microbubbles and larger bubbles in five regions were calculated. A volume of 10 microL of 47.5% ethanol was effective in defoaming larger bubbles generated by the SM test procedure without altering the stable microbubble counts. When concurrently performed on 43 samples of gastric aspirate obtained at birth from infants of less than 35 weeks gestation, the RDS predictive value of the ERM test was similar to that of the SM test. It was concluded that the ERM test may serve as an alternative to the SM test.

Effect of Calcium Chloride on Sodium Alginate Microbubbles as Ultrasound Contrast Agent

In order to stabilize microbubbles without coating layers to be used as ultrasound contrast agent, the viscosity of the liquid for storage has to be increased. Sodium alginate is a material which forms viscous liquid when its powder is dissolved in water. The viscosity may vary from 1 cP to about 100 cP when the amount of material used is increased from 0.1% to 1%. However, the viscosity is still not enough to make stable microbubbles which can be stored for over several weeks. In this paper, by adding calcium chloride to the sodium alginate solution, a gel-type solution is produced which has a high viscosity capable of holding microbubbles for a long time. By measuring the viscosity of the sodium alginate solution and the solution with calcium chloride solution added, it is known that the solution is changed from a Newtonian liquid to a non-Newtonian liquid when calcium chloride solution is added. Since the attenuation coefficient of microbubble dispersion is proportional to the number of microbubbles, the disappearance of microbubbles lowers the attenuation coefficient. Therefore, in this study, the attenuation coefficient is used as an estimating parameter for the lifetime of microbubbles.

Langmuir Trough Study of Surfactant Mixtures Used in the Production of a New Ultrasound Contrast Agent Consisting of Stabilized Microbubbles

Span-type surfactants (sorbitan fatty acid esters) and Tween-type surfactants (sorbitan polyoxyethylene fatty acid esters) are employed by us to generate stabilized microbubbles for use in diagnostic ultrasound. After sonication of an aqueous surfactant solution, only some mixtures of Span-type and Tween-type surfactants at certain conditions can form stable microbubbles. This work investigated the stability of the surfactant-stabilized microbubbles by using a Langmuir trough to measure the π−A isotherms of the surfactant monolayer. The experimental results, which agreed with a theoretical analysis of the microbubble stability, indicate that the surfactant-stabilized microbubbles have a solid-condensed monolayer “skin” which functions to reduce the surface tension, prevent coalescence between microbubbles, and increase their aqueous compatibility. The higher surface pressure obtained for the case of a microbubble preparation, compared with that of unsonicated mixtures, indicates that sonication enhances t...

Structural studies on stabilized microbubbles: development of a novel contrast agent for diagnostic ultrasound

Ultrasound is a noninvasive tool with which the physician can gain insight into the state of the internal organs of the body. A contrast agent is a substance which, when injected into the body improves the resolution of an image. The use of diagnostic ultrasound is limited without development of such an agent. This paper describes the development of a new class of contrast agent. The agent consists of microbubbles generated in a solution by sonication, and stabilized by a layer of nonionic surfactant molecules. A single surfactant type alone does not produce stable bubbles, and only certain combinations of surfactants, one with a hydrophile-lipophile balance (HLB) greater than 10.5 and the other with an HLB less than 9, are successful. An agent prepared from Span60® and Tween80® is described in detail. A Coulter analysis revealed that 90% of the bubbles were less than 10 μm in diameter. This is essential if the agent is to pass the pulmonary capillary bed. B-mode imaging of a sample of microbubbles indicated that these bubbles were highly echogenic (that is they produced a strong contrast). A Langmuir trough study of the molecular arrangement of the surfactant molecules inside the microbubble skin suggested that there are 1.7 molecules of Span to each Tween molecule. Detailed analysis of the pressure-area curves and area per molecule data lead to the hypothesis that the correct proportion of Span molecules can substantially reduce the head group repulsion found in the all-Tween situation, and this results in the generation of stable microbubbles. A molecular arrangement of these surfactant molecules around the microbubble is proposed.

Ultrasound and microbubbles: Their generation, detection and potential utilization in tissue and organ therapy—Experimental

Ultrasound-induced cavitation in tissue and organs has been well recognized and documented. Generally, this phenomenon has been seen as something to be avoided except in cases such as lithotripsy, where its production is considered an essential part of the treatment process or as a desirable contrast media in some areas of visualization enhancement. This article covers theree areas in which the phenomenon has been observed, and shows how the effect can or may be therapeutically beneficial. Studies in the pig show that implanted human gallstones and the gallbladder itself can be eliminated in a nonsurgical procedure using ultrasound-induced cavitation in the gallbladder. In the dog brain, relatively stable cavitation-induced microbubbles have been transported through the vascular system to regions outside a focal seeding site. These bubbles produce ablation of tissue volumes at a remote site when irradiated with appropriate ultrasound. The cavitation phenomenon has been observed in the dog and human prostate. In the human prostate, microbubbles transported from ultrasound-induced focal seeding sites can be readily visualized with ultrasound and may be potentially useful under controlled conditions in tissue debulking for the treatment of benign prostatic hyperplasia (BPH). A similar microbubble transport has not been seen in the dog prostate under similar ultrasound treatment parameters. The ability to detect cavitation-induced microbubbles, follow their transportation through the vascular system and excite them at the appropriate time and place provides interesting possibilities for therapy. Of course, the entire microbubble process can be avoided by working below the cavitation threshold, thereby using only the absorption of ultrasound in tissue to produce focal thermal lesions. The term microbubble is used here in the context of those bubbles which can be transported in the vascular system down to vessels diameters below the 100-μm range. This is the vessel size in the vascular field into which microbubbles are transported and can be both visualized as well as disrupted with ultrasound.

5215680 Method for the production of medicalgrade lipid-coated microbubbles, paramagnetic labeling of such microbubbles and therapeutic uses of microbubbles

This invention relates to a large scale method for the production of medical grade lipid-coated microbubbles, to the paramagnetic labeling of such microbubbles and to therapeutic applications for the microbubbles. More particularly, the invention relates to a method of the production of medical grade, concentrated suspensions of stable, paramagnetically derivatized or underivatized microbubbles useful for ultrasonic and magnetic resonance imaging and also relates to therapeutic interventions such as selective tumor destruction.

A transpulmonary contrast medium enhances the transcranial Doppler signal in humans.

Transtemporal insonation in transcranial Doppler sonography is often impaired by an insufficient signal-to-noise ratio, especially in elderly patients. A transpulmonary stable air microbubble suspension was injected intravenously in humans as an intracranial ultrasonic contrast agent. In a clinical phase II study, 20 patients (15 women, 5 men; mean age, 65.5 +/- 11.5 years) presenting with clinical indications for transcranial Doppler investigation were examined. A total of 97 intravenous injections with different concentrations (200, 300, and 400 mg/mL of suspension) of air microbubbles bound to galactose microparticles as a carrier were performed. The signal enhancement of color-coded pulse curves of basal cerebral arteries was evaluated off-line in comparison to an integrated color-coded decibel scale, considering quality, quantity, and time course of enhancement requiring a 3-dB level above the native signal. The overall diagnostic information was assessed according to a reliability score. The first acoustic signal increase was registered after an average of 21 seconds. Time intervals for a dose-dependent peak intensity and maximal duration were 41.3 +/- 17.1 seconds and 118.0 +/- 69.8 seconds (200 mg/mL); 55.5 +/- 27.7 seconds and 237.0 +/- 112.3 seconds (300 mg/mL); and 66.1 +/- 31.8 seconds and 293.0 +/- 122.0 seconds (400 mg/mL), respectively. Duration of signal enhancement increased significantly (P < .05) with higher concentrations. The extent of signal enhancement during the whole pulse curve reached an average of 9.1 +/- 5.0 dB for 200 mg/mL, 12.0 +/- 5.4 dB for 300 mg/mL (significant on P < .05 level), and 13.1 +/- 5.6 dB for 400 mg/mL concentration (P = NS). Respective maximal intensity spots reached 17.5 +/- 6.0, 20.7 +/- 5.5, and 22.7 +/- 5.9 dB for increasing concentrations, respectively. Overall visual assessment of enhanced pulse curves for diagnostic reliability showed a sufficient result in 38.1% of all injections with 200 mg/mL, in 88.6% with 300 mg/mL, and in 84.2% with 400 mg/mL concentration. Minimal side effects occurring in 12.4% of all injections were all reversible. Transpulmonary stable air microbubbles bound to a galactose carrier represent a useful and safe contrast agent in case of an insufficient native signal in transcranial Doppler investigation.

Galactose-based intravenous sonographic contrast agent: experimental studies.

A galactose-based sonographic contrast agent, which produces stable microbubbles capable of traversing the cardiopulmonary circulation, was used to enhance Doppler signals in blood vessels of varying size after intravenous injection. A series of experiments using dogs, rabbits, and woodchucks was conducted to establish the ability of the agent to enhance the reflectivity of normal tissue, tumor tissue, and blood. Although no enhancement was perceptible in tissue on the sonogram, significant enhancement of color and spectral Doppler signals was demonstrated in a variety of vessels. These included the aorta, vena cava, and portal vein as well as such small vessels as those of the retina of the eye, renal cortex, liver parenchyma, and gallbladder wall. Both spectral and color Doppler enhancement was shown in naturally occurring woodchuck hepatomas. Peak Doppler signal enhancement after bolus injection was approximately 10 dB with a dose of 0.01 ml/kg. Recirculation of the agent provided enhancement after intravenous bolus injection for more than 3 min. With a steady intravenous infusion of 0.2 ml/min/kg, Doppler signal enhancement of about 14 dB was maintained continuously for more than 5 min. The results of these animal experiments, in particular in small vessels and with recirculation after intravenous injection, suggest excellent potential for future clinical applications.

The acoustic filter: An ultrasonic blood filter for the heart-lung machine

Cardiopulmonary bypass-associated encephalopathy is thought to be due in part to continuous microembolization of the brain with gas microbubbles more than 40 microns in diameter during bypass. Current barrier filter technology cannot effectively remove such small microbubbles in fragile fluids such as blood. The design concepts for a new nonbarrier ultrasound-based fluid filtration system (an "acoustic filter") capable of filtering small microbubbles from blood are presented. The acoustic filter uses a field of high-intensity ultrasound to push microbubbles down an acoustic gradient, where they can be collected and removed. To test the filtration efficiency of the system, a Doppler ultrasound bubble detector was built. By monitoring the prefilter and postfilter Doppler signal an assessment of filtration efficiency was made. A suspension of stable albumin-encapsulated microbubbles (4 to 32 microns) were used as a model of the microbubble contaminants that might be found in the arterial return line of the heart-lung machine. Inactivated, the acoustic filter neither removed nor added microbubbles to the fluid. Activated, the acoustic filter provided total or near-total clearing of microbubbles. We conclude that the acoustic filter can remove microbubbles from a cardiopulmonary bypass-like apparatus.

Seamount acoustic scattering

The cover of the March 1 issue of Eos showed a time series of acoustic scattering above Southeast Hancock Seamount (29°48′N, 178°05′E) on July 17–18, 1984. In a comment on that cover Martin Hovland (Eos, August 2, p. 760) argued that gas or “other far reaching causes” may be involved in the observed acoustic signals. He favors a hypothesis that acoustic scattering observed above a seeping pockmark in the North Sea is a combination of bubbles, stable microbubbles, and pelagic organisms and infers that this may be a more general phenomenon and indeed plays a role in the attraction of organisms to seamounts

Microbubbles: Stabilization by monolayers of adsorbed particles

Stable microbubbles are a ubiquitous feature of natural waters, acting as sites for mechanical and acoustical cavitation and nucleii for bubble growth at low gas super‐saturations. Stable microbubble populations that have been determined acoustically in the past are not well explained by existing models of bubble stabilization, i.e., stabilization in particle crevices or stabilization by molecular monolayers. Based upon the observation that bubbles rising in sea water rapidly become coated with particles, a model is developed in which bubbles are stabilized mechanically by monolayers of adsorbed nonpolar particles. Long‐term bubble stability is demonstrated in laboratory experiments. Predictions based on the experiments and the model are, in order of magnitude, consistent with observation.

Two-dimensional contrast echocardiography. I. In vitro development and quantitative analysis of echo contrast agents

To facilitate the passage of echo contrast agents through the microcirculation and the echocardiographic study of myocardial perfusion, ultrasonic energy (sonication) was employed to produce contrast agents consisting of relatively uniform, stable and small (less than 10 mu diameter) gaseous microbubbles suspended in liquid solutions. The size and persistence of the microbubbles was verified by light microscopy and an in vitro system were employed for comparative assessment of peak echo amplitude and echo persistence characteristics of various contrast agents. The study indicated that although a variety of hand-agitated and sonicated contrast agents provided satisfactory echo intensities, sonication was clearly superior to the hand-agitation method, because sonication produced smaller, more uniform and more stable microbubbles that may be suitable for myocardial contrast echocardiography. It is concluded that of the contrast agents examined, sonicated solutions of sorbitol (70%) and dextrose (70%) appeared to have particular potential because of the small sizes of the microbubbles (6 +/- 2 and 8 +/- 3 mu, respectively) and their prolonged in vitro persistence. The use of sonication to produce standardized, small and stable microbubbles should facilitate physiologic passage of the contrast agent through the capillary beds and allow two-dimensional imaging of the left heart myocardium during right-sided, aortic root, coronary sinus or intracoronary contrast injections.

Echogenicity caused by stable microbubbles in a protein-lipid emulsion.

AbstractSome investigators have hypothesized that the echogenicity of natural emulsions is due to protein‐lipid interfaces. An in vitro investigation of these phenomena indicates that the observed echoes are the result of entrapped microbubbles that can remain relativelystable in certain naturally occurring products, such as buttermilk.

Maturity of fetal lungs tested by production of stable microbubbles in amniotic fluid.

Lung surfactant in amniotic fluid, and hence the maturity of the fetal lungs, can be assessed by observation of stable microbubbles (less than 15 micron diameter). Bubbles are formed by agitation with a Pasteur pipette and examined in hanging drops under the 10 x power of a microscope. Either after a count of bubbles, or after a general survey of hanging drops, the fluid is given a stable microbubble rating. A 'strong' rating indicates that the idiopathic respiratory distress syndrome will not occur after delivery, and that the lecithin/sphingomyelin ratio will indicate maturity. Complete absence of stable microbubbles suggests a high risk of respiratory trouble for the newborn infant, as does a weak or lower rating in the 30 to 37 week gestational age group. The test takes 5 to 10 minutes to perform, is cheap and easy, is not affected by blood, but may be affected by meconium. If a 'strong' rating is found, measurement of the L/S ratio can safely be omitted.