Near-Infrared Fluorescence Imaging Contrast Agents for Clinical Research: Limitations and Alternatives

Introduction: Near-infrared fluorescence imaging is a technique that will establish itself in the short term at the international level because it is recognized for its potential to improve the performance of surgical interventions, its moderate investment and operating costs and its portability. Although the technology is now mature, there is currently the problem of the availability of contrast agents to be injected IV. The aim of this methodology article is to propose an alternative solution to the need for contrast agents for clinical research, particularly in oncology. Methodology: They consist of coupling a fluorescent marker in the form of an NHS derivative, such as IR DYE manufactured in compliance with GMP, with therapeutic monoclonal antibodies having marketing authorization for molecular imaging. For a given antibody, the marking procedure must be the subject of a validation file on the final preparation filtered on a sterilizing membrane at 0.22 μm. Once the procedure has been validated, it would be unnecessary to repeat the tests before each clinical research examination. A check of the marking by thin-layer chromatography (TLC) and place it in a sample bank at +4˚C for 1 month of each injected formulation would be sufficient for additional tests if necessary. Conclusion: Molecular near-infrared fluorescence imaging is experiencing development, the process of which could be accelerated by greater availability of clinical contrast agents. Alternative solutions are therefore necessary to promote clinical research in this area. These methods must be shared to make it easier for researchers.

Application of Low Tube Voltage, Low-concentration Contrast Agent Using a 320-row CT in Coronary CT Angiography: Evaluation of Image Quality, Radiation Dose and Iodine Intake

The effect of low voltage and low concentration contrast agent on image quality of coronary CT angiography,radiation dose and iodine intake was evaluated.A total of 121 patients with body mass index(BMI)<26 kg/m2 and heart rate(HR)<70 beats/min were randomly divided into four groups:group A(n=31,80 kVp,270 mgl/mL);group B(n=33,100 kVp,270 mgl/mL);group C(h=30,100 kVp,320 mgl/mL);group D(w=27,100 kVp,400 mgl/mL).The automatic current modulation system and the iterative algorithm for reconstruction were adopted in each group.The CT values and SD values of the aortic root(AR),subcutaneous fat,left coronary artery opening(LCA),and right coronary artery opening(RCA)were measured in all groups,the signalto-noise ratio(SNR)and contrast noise ratio(CNR)were calculated,and effective radiation dose and iodine intake were recorded.The subjective assessment for image quality was performed by two physicians using a 4-point scale.The results were compared using the one-way ANOVA and rank sum tests.The image quality of the four groups met the clinical diagnostic requirements.The CT values of AR in groups A,B,C,and D were 537.6±71.4,447.2±81.9,445.2±64.9 and 518.5±94.9 Hu,respectively,with no significant difference between group A and group D,or between group B and group C,while CT values in groups B and C were significantly lower than those in groups A and D(P<0.05).In groups A,B,C,and D,the LCA SNR values were 22.7±9.1,23.3±9.1,23.3±7.7 and 26.6±8.9,and the RCA CNR values were 26.9±9.&28.5±11.4,27.7土&8 and 32」±10.6,respectively.The AR visual scores in groups A,B,C and D were 3.8±0.2,3.9±0.3,3.9±0.3 and 4.0±0.3,respectively.There were no significant differences in SNR,CNR and visual score among the four groups(P>0.05).The radiation doses in groups A,B,C and D were 2.6±1.4,3.6±1.&4.9±3.5 and 4.9±2.8 mSv,respectively.The radiation dose in group A was significantly less than that in the rest three groups(P<0.05).The iodine intakes in groups A,B,C and D were 14.9±1.5,15.0±1.5,17.7±2.0 and 18.1±2.5 g,respectively.There was no significant difference in the intake of iodine between groups C and D,or between groups A and B,while iodine intake in groups A and B were significantly reduced as compared with that in groups C and D(P<0.05).It was concluded that for patients with low BMI and controlled HR,compared to 100 kVp tube voltage combined with multiple concentration contrast agents,80 kVp combined with 270 mgl/mL contrast agent is enough to ensure the quality of the images,and can reduce the radiation dose significantly,while reducing the amount of iodine intake notably,thus reducing the incidence of adverse reaction.

Non-thermal ablation of rabbit liver VX2 tumor by pulsed high intensity focused ultrasound with ultrasound contrast agent:Pathological characteristics

AIM： To investigate the pathological characteristics of non-thermal damage induced by pulsed high intensity focused ultrasound （PHIFU） combined with ultrasound contrast agent （UCA）, SonoVue （Bracco SpA, Milan, Italy） in rabbit liver VX2 tumor. METHODS： Liver VX2 tumor models were established in 20 rabbits, which were divided randomly into PHIFU combined with ultrasound contrast agent group （PHIFU ＋ UCA group） and sham group. In the PHIFU ＋ UCA group, 0.2 mL of SonoVue was injected intravenously into the tumor, followed by ultrasound exposure of Isp 5900 W/cm^2. The rabbits were sacrificed one day after ultrasound exposure. Specimens of the exposed tumor tissues were obtained and observed pathologically under light microscope and transmission electron microscope. The remaining tumor tissues were sent for 2,3,5-Triphenyltetrazolium chloride （TTC） staining. RESULTS： Before Trc staining, tumor tissues in both the sham and the PHIFU ＋ UCA groups resembled gray fish meat, After TIC staining, the tumor tissues were uniformly stained red, with a clear boundary between tumor tissue and normal tissue, Histological examination showed signs of tumor cell injury in PHIFU ＋ UCA group, with cytoplasmic vacuoles of various sizes, chromatin margination and karyopyknosis. Electron microscopic examination revealed tumor cell volume reduction, karyopyknosis, chromatin margination, intercellular space widening, the presence of high electro＇n-density apoptotic bodies and vacuoles in cytoplasm. CONCLUSION： The non-thermal effects of PHIFU combined with UCA can be used to ablate rabbit liver VX2 tumors.

Contrast-enhanced ultrasound using SonoVue mixed with oral gastrointestinal contrast agent to evaluate esophageal hiatal hernia: Report of three cases and a literature review

BACKGROUND Due to a thicker abdominal wall in some patients,ultrasound artifacts from gastrointestinal gas and surrounding tissues can interfere with routine ultrasound examination,precluding its ability to display or clearly show the structure of a hernial sac(HS)and thereby diminishing diagnostic performance for esophageal hiatal hernia(EHH).Contrast-enhanced ultrasound(CEUS)imaging using an oral agent mixture allows for clear and intuitive identification of an EHH sac and dynamic observation of esophageal reflux.CASE SUMMARY In this case series,we report three patients with clinically-suspected EHH,including two females and one male with an average age of 67.3±16.4 years.CEUS was administered with an oral agent mixture(microbubble-based SonoVue and gastrointestinal contrast agent)and identified a direct sign of supradiaphragmatic HS(containing the hyperechoic agent)and indirect signs[e.g.,widening of esophageal hiatus,hyperechoic mixture agent continuously or intermittently reflux flowing back and forth from the stomach into the supradiaphragmatic HS,and esophagus-gastric echo ring(i.e.,the“EG”ring)seen above the diaphragm].All three cases received a definitive diagnosis of EHH by esophageal manometry and gastroscopy.Two lesions resolved upon drug treatment and one required surgery.The recurrence rate in follow-up was 0%.The data from these cases suggest that the new non-invasive examination method may greatly improve the diagnosis of EHH.CONCLUSION CEUS with the oral agent mixture can facilitate clear and intuitive identification of HS and dynamic observation of esophageal reflux.

Self-assembled superparamagnetic nanoparticles as MRI contrast agents — A review

Recent progress of the preparation and applications of superparamagnetic iron oxide（SPIO） clusters as magnetic resonance imaging（MRI） probes is reviewed with regard to their applications in labeling and tracking cells in vivo, in diagnosis of cardiovascular diseases and tumors, and in drug delivery systems. Magnetic nanoparticles（NPs）, especially SPIO nanoparticles, have long been used as MRI contrast agents and as an advantageous nanoplatform for drug delivery,taking advantage of their unique magnetic properties and ability to function at the molecular and cellular levels. Due to advances in nanotechnology, various means to control SPIO NPs＇ size, composition, magnetization and relaxivity have been developed, as well as ways to usefully modify their surface. Recently, self-assembly of SPIO NP clusters in particulate carriers — such as polymeric micelles, vesicles, liposomes, and layer-by-layer（Lb L） capsules — have been widely studied for application as ultrasensitive MRI probes, owing to their remarkably high spin–spin（T2） relaxivity and convenience for further functionalization.

Photoacoustic molecular imaging using combined acupuncture and gold nanorods as a composite contrast agent

In this study,we developed a novel photoacoustic imaging technique based on poly(ethyleneglycol)-coated(PEGylated)gold nanorods(PEG-GNRs)(as the contrast agent)combined with traditional Chinese medicine(TCM)acupuncture(as the auxiliary method)for quantitatively monitoring contrast enhancement in the vasculature of a mouse brain in vivo.This study takes advantage of the strong near-infrared absorption(peak at700 nm)of GNRs and the ability to adjust the hemodynamics of acupuncture.Experimental results show that photoacoustic tomography(PAT)successfully reveals the optical absorption variation of the vasculature of the mouse brain in response to intravenous administration of GNRs and acupuncture at the Zusanli acupoint(ST36)both individually and combined.The quantitative measurement of contrast enhancement indicates that the composite contrast agents(integration of acupuncture and GNRs)would greatly enhance the photoacoustic imaging contrast.The quantitative results also have the potential to estimate the local concentration of GNRs and even the real-time effects of acupuncture.

Oleic Acid (OA), A Potential Dual Contrast Agent for Postmortem MR Angiography (PMMRA): A Pilot Study

Summary:Choosing proper perfusates as contrast agents is an important aspect for postmortem magnetic resonance angiography(PMMRA).However,in this emerging field,the number of suitable kinds of liquid is still very limited.The objective of this research is to compare MR images of oleic acid(OA)with paraffin oil(PO)in vitro and in ex situ animal hearts,in order to evaluate the feasibility to use OA as a novel contrast agent for PMMRA.In vitro,OA,PO and water(control)were introduced into three tubes separately and T,weighted-spin echo(Tw SE)and T2w-SE images were acquired on a 1.5T MR scanner.In the second experiment,0A and PO were injected into left coronary artery(LCA)and left ventricle(LV)of ex situ bovine hearts and their Tw-SE,Tzw-SE,Tw-multipoint Dixon(Tjw-mDixon)and 3DT2w-mDixon images were acquired.The overall results indicate that OA may have a potential to be used as a dual(T and T2 based)contrast agent for PMMRA when proper sequence parameters are utilized.However,as the pilot study was based on limited number of animal hearts,more researches using OA in cadavers are needed to validate our findings.

Role of microbubble ultrasound contrast agents in the non-invasive assessment of chronic hepatitis C-related liver disease

Patients who are chronically infected with the hepatitis C virus often develop chronic liver disease and assessment of the severity of liver injury is required prior to considering viral eradication therapy. This article examines the various assessment methods currently available from gold standard liver biopsy to serological markers and imaging. Ultrasound is one of the most widely used imaging modalities in clinical practice and is already a first-line diagnostic tool for liver disease. Microbubble ultrasound contrast agents allow higher resolution images to be obtained and functional assessments of microvascular change to be carried out. The role of these agents in quantifying the state of hepatic injury is discussed as a viable method of determining the stage and grade of liver disease in patients with hepatitis C. Although currently confined to specialist centres, the availability of microbubble contrast-enhanced ultrasound will inevitably increase in the clinical setting.

Study on Multi-density Contrast Agent Fillers of Duct Casting Based on CT Three-Dimensional Reconstruction

The three-dimensional visualization model of human body duct is based on virtual anatomical structure reconstruction with duct angiography,which realizes virtual model transferred from two-dimensional,planar and static images into three-dimensional,stereoscopic and dynamic ones repectively.In recent years,the multi-duct segmentation and division of the same specimen（or organ） is the focus of attention shared by surgeons and clinical anatomists.On the basis of 4.22 g/cm3 body bone density,this study has screened out metal oxide contract agent with different density for infusion and modeling,as well as compared and analyzed the effects of three-dimensional image of CT virtual bronchoscopy（CTVB）,three-dimensional image of CT maximum intensity projection and three-dimensional model.This experiment result showed synchronously infusing multi-duct of same specimen（or organ） with contrast agent in different densities could reconstruct three-dimensional models of all ducts once only and adjust threshold to develop single or multiple ducts.It was easier to segment and observe the duct structure,anastomosis,directions and crossing in different parts,which was beyond comparison with three-dimensional image of CTVB.Although the existing three-dimensional duct reconstruction techniques still cannot be applied in living bodies temporarily,this study focused on a creative design of ducts segmentation in different density,which proposed a new experimental idea for developing multi-duct three-dimensional model in living body in the future.It will play a significant role in disease diagnosis and individual design in surgical treatment program.Therefore,this study observes the three-dimensional status of human duct with the application of contrast agent fillers in different density,combined with three-dimensional reconstruction technology.It provides an innovative idea and method for constructing three-dimensional model of digital multi-duct specimen,and the ultimate goal is to develop the digitized virtual human and precise medical treatment better and faster.

PREPARATION OF POLYELECTROLYTE MULTILAYER COATED MICROBUBBLES FOR USE AS ULTRASOUND CONTRAST AGENT

Objeelive To prepare and characterize polyelectrolyte multilayer film coated microbubbles for use as ultrasound contrast agent （UCA） and evaluate its effects in ultrasonic imaging on normal rabbit＇s fiver parenchyma. Methods Perfluorocarbon （PFC）-containing microbubbles （ST68-PFC） were prepared by sonication based on suffactant （ Span 60 and Tween 80）. Subsequently, the resulting ST68-PFC microbubbles were coated using oppositely charged polyelectrolytes by microbubble-templated layer-by-layer self-assembly technique via electrostatic interaction. The enhancement effects in ultrasonic imaging on normal rabbit＇s liver parenchyma were assessed. Results The obtained microbubbles exhibited a narrow size distribution. The polyelectrolytes were successfully assembled onto the surface of ST68-PFC microbubbles. In vivo experiment showed that polyelectrolyte multilayer film coated UCA effectively enhanced the imaging of rabbit＇s liver parenchyma. Conclusions The novel microbubbles UCA coated with polyelectrolyte multilayer, when enabled more function, has no obvious difference in enhancement effects compared with the pre-modified microbubbles. The polymers with chemically active groups （ such as amino group and carboxyl group） can be used as the outermost layer for attachment of targeting ligands onto microbubbles, allowing selective targeting of the microbubbles to combine with desired sites.

Nanomaterials incorporated ultrasound contrast agents for cancer theranostics

Nanotechnology provides various nanomaterials with tremendous functionalities for cancer diagnostics and therapeutics.Recently, theranostics has been developed as an alternative strategy for efficient cancer treatment through combination of imaging diagnosis and therapeutic interventions under the guidance of diagnostic results. Ultrasound(US) imaging shows unique advantages with excellent features of real-time imaging, low cost, high safety and portability, making US contrast agents(UCAs)an ideal platform for construction of cancer theranostic agents. This review focuses on the development of nanomaterials incorporated multifunctional UCAs serving as theranostic agents for cancer diagnostics and therapeutics, via conjugation of superparamagnetic iron oxide nanoparticles(SPIOs), Cu S nanoparticles, DNA, si RNA, gold nanoparticles(GNPs), gold nanorods(GNRs), gold nanoshell(GNS), graphene oxides(GOs), polypyrrole(PPy) nanocapsules, Prussian blue(PB) nanoparticles and so on to different types of UCAs. The cancer treatment could be more effectively and accurately carried out under the guidance and monitoring with the help of the achieved theranostic agents. Furthermore, nanomaterials incorporated theranostic agents based on UCAs can be designed and constructed by demand for personalized and accurate treatment of cancer, demonstrating their great potential to address the challenges of cancer heterogeneity and adaptation, which can provide alternative strategies for cancer diagnosis and therapeutics.

Predictive Study of Velocimetry in the Coronary Artery after Iodinated Contrast Agent Injection

This study aims to determine the retention time of iodinated contrast agents (ICA) in the coronary artery. The mechanical aspect of ICA displacement was studied in order to better understand the effect of these products on the inner wall of the artery of patients with coronary artery stenosis, undergoing repeated imaging examinations with iodinated contrast agents. ICA flow, ICA and blood flow in the artery were modelled. The fluid was regarded to be viscous, incompressible and Newtonian. Blood flow was presumed to be unidirectional, laminar and unstationary. Iodine flow velocity and retention time were calculated using the Runge-Kutta 4th order method programmed in C++ and MatLab R2013a language. The results showed that for coronary artery CT-scans, ICA retention time is 1 minute 40 seconds and for coronary arteriography it is between 2.41 and 3.61 seconds. The values calculated were compared to theoretical values and to clinical observations. The results enabled us to validate our model.

Combined Pluronic P85- and Ultrasound Contrast Agents-mediated Gene Transfection to HepG2 Cells

This study examined the effect of P85 （a pluronic block copolymer） and microbubble （MB） ultrasound contrast agents under ultrasound irradiation on gene transfection and expression. The pEGFP plasmids that can encode enhanced green fluorescent protein （pEGFP） served as a report gene and were mixed with different concentrations of MB/0.05% （w/v） P85. Then the plasmids were transfected into human hepatoma G2 （HepG2） cells. The HepG2 cells treated with MB/P85 or without treatment were exposed to ultrasound （US parameters： 1 MHz, 1.0 W/cm2, 20 s, 20% duty cycle）. Twenty-four hours later, the transfection efficiency was assessed by fluorescence microscopy and fluo-rescence activated cell sorting （FACS） analysis. The cell viability was evaluated by Trypan blue exclusion test. The results showed that the gene transfection efficiency in HepG2 cells under ultrasound irradiation was significantly higher than that without ultrasound irradiation. HepG2 cells in the MB or P85 group in the absence of ultrasound expressed less amount of green fluorescent protein. The expression efficiency reached （22.14±3.06）% and the survival rate was as high as （55.73±3.32）% in the 30% MB plus P85 group. It was concluded that MB and P85 in the presence of ultrasound can enhance gene transfection and expression.

Dynamic Assessment of the Focal Hepatic Lesion in Rats Using Ultrasonic Contrast Agent

The focal hepatic lesion caused by local injection of absolute alcohol in rats was evaluated with ultrasonic contrast agent and pathologic examination. Twenty adult Wistar rats weighing about 200 g were injected with absolute alcohol （0. 05-0. 1 mL each one） on the exterior left lobe of the liver under the monitoring of ultrasound. Pulse inversion harmonic imaging was used to evaluate the focal lesion after bolus injection of ultrasonic contrast agent （0. 05 mL/200 g） through caudal vein. Seven days later, the focal lesion was studied again as before. The exterior left lobe of liver with focal lesion was incised and underwent pathologic examination. The results showed that all of the focal lesions could be defined clearly after bolus injection of the ultrasonic contrast agent under the mode of pulse inversion harmonic imaging. There was good correlation between the size of the focal lesion measured by ultrasound on the 7th day after the ＂ablation＂ under the mode of pulse inversion harmonic imaging and that gotten by pathologic examination （P=0.39）. The focus size measured by ultrasound right after the ablation was larger than that gotten by pathologic examination （P=0. 002）. It was concluded that ultrasonic contrast agent plus pulse inversion harmonic imaging could be used to assess the size of the focal hepatic lesion caused by local injection of absolute alcohol in rats.

Lactoferrin-Conjugated Polylactic Acid Nanobubbles Encapsulated Perfluoropentane as a Contrast Agent for Ultrasound/Magnetic Resonance Dual-Modality Imaging

The development of contrast agents that can be activated by multiple modes is of great significance for tumor diagnosis.In this study,the lactoferrin(Lf)-conjugated polylactic acid(PLLA)nanobubbles(Lf-PLLA NBs)were used to encapsulate liquid perfluoropentane(PFP)with the double emulsion method,creating PFP loaded(PFP/Lf-PLLA)NBs for the ultrasound/magnetic resonance dual-modality imaging of subcutaneous tumor.The parti-cle diameter and stability of nanobubbles were investigated by photon correlation spectroscopy.The biocompat-ibility of nanobubbles was preliminarily evaluated by cell proliferation and migration assay,hemolysis rate,and blood biochemistry analysis.A B-mode clinical ultrasound real-time imaging system was used to perform ultra-sonic imaging in vivo.Magnetic resonance imaging in vivo was applied with a clinical 3.0 T magnetic resonance imaging(MRI)scanner system.The mean particle diameter of PFP/Lf-PLLA NBs was 320.2±4.1 nm with a low polydispersity index(PDI,0.145±0.025),and the NBs were negatively charged(−11.4±0.4 mV).The transmis-sion electron microscopy(TEM)results showed that PFP/Lf-PLLA NBs exhibited highly monodispersed and pos-sessed an obvious spherical structure of nanocapsules.Nanobubbles had good stability at 4°C.Different concentrations of the PFP/Lf-PLLA NBs solution had no effect on the cell in cytotoxicity and cell migration,and the results of hemolysis rate and blood biochemistry assay also indicated the good biocompatibility of NBs.On the ultrasound/magnetic resonance imaging of tumor-bearing mice,PFP/Lf-PLLA NBs showed signifi-cantly enhanced contrast ability of tumor tissue.Therefore,PFP/Lf-PLLA NBs had great potential to be a contrast agent for tumor dual-modality imaging in vivo.

Mesoporous MnSiO3@Fe3O4@C Nanoparticle as p H-responsive T1-T2* Dual-modal Magnetic Resonance Imaging Contrast Agent for Tumor Diagnosis

Mesoporous structured MnSiO3@Fe3O4@C nanoparticles(NPs)were prepared via a facile and efficient strategy,with negligible cytotoxicity and minor side efforts.The as-prepared MnSiO3@Fe3O4@C NPs hold great potential in serving as pH-responsive T1-T2^*dual-modal magnetic resonance(MR)imaging contrast agents.The released Mn^2+shortened T1 relaxation time,meanwhile the superparamagnetic Fe3O4 enhanced T2 contrast imaging.The release rate of Mn ions reaches 31.66%under the condition of pH=5.0,which is similar to tumor microenvironment and organelles.Cytotoxicity assays show that MnSiO3@Fe3O4@C NPs have minor toxicity,even at high concentrations.After intravenous injection of MnSiO3@Fe3O4@C NPs,a rapid contrast enhancement in tumors was achieved with a significant enhancement of 132%after 24 h of the administration.Moreover,a significant decreasement of 53.8%was witnessed in T2 MR imaging signal.It demonstrated that MnSiO3@Fe3O4@C NPs can act as both positive and negative MR imaging contrast agents.Besides,owing to the pH-responsive degradation of mesoporous MnSiO3,MnSiO3@Fe3O4@C NPs can also be used as potential drug systems for cancer theranostics.

Oral Gd-DTPA as a negative gastrointestinal contrast agent in magnetic resonance cholangiopancreatography

Objective: To evaluate the value of oral Gd-DTPA as a negative contrast agent during magnetic resonance cholangiopancreatography (MRCP) to eliminate the high signals of the gastrointestinal tract. Methods: To select the optimal concentration of oral Gd-DTPA for MRCP, a phantom study was performed followed by clinical trial in 15 cases undergoing MRCP before and after oral Gd-DTPA (in a total volume of 250 ml 1∶5 diluted Gd-DTPA, 1.488 g/L). MRCP images were acquired using two-dimensional single slice fast spin-echo (SSTSE) sequence and half-Fourier acquisition single slice fast spin-echo (HASTE) sequence. Results: The phantom study showed that the 1∶5 diluted oral Gd-DTPA was best in decreasing the signal intensity both in T2-weighted imaging (59.5%) and in HASTE sequence (82.45%). The high signal intensity of the stomach and intestinal fluid was completely suppressed in all the cases. The depictions of the common bile duct and pancreatic duct were markedly improved by using the oral contrast agent (P<0.05). Conclusion: Oral Gd-DTPA is effective and safe for eliminating the high signal of the gastrointestinal tract to improve the depiction of the biliary system by MRCP.

Microbubble ultrasound contrast agent with three esters and carboxylic methyl cellulose as main shell materials: Its preparation and imaging evaluation

Objective: To study on the preparation process of a new surfactant-based microbubble ultrasound contrast agent and to evaluate its contrast effects in vivo. Methods: Microbubble ultrasound contrast agent with three ester surfactants and other additives as its shell materials was prepared by sonication. Sulfur hexafluoride was adopted as the inner gas of the microbubbles. New methods through the combination of optical microscope and some softwares were used to measure the size distribution and the concentration of the microbubbles. Some parameters such as the pH value of the phosphate buffer, quantity of the carboxylic methyl cellulose in the shell materials, selection of the ultrasound power and process time, were studied. Six hybirded dogs were used to verify the in vivo contrast imaging of the contrast agent using second harmonic power Doppler modality. Safety and persistent time of the agent inner animal body were also investigated. Results: Ultrasound contrast agent prepared in the experiment had an average microbubble diameter of 3.95 microns with concentration of 3.6×109 microbubbles per millilitre. Carboxylic methyl cellulose was found as an important shell material which had obviously effect on the microbubble stability and production even with a little quantity. The buffer pH value also had a key role on the microbubble formation and the final production. When the buffer pH value reached 7.4, there was no microbubble produced. Under the approximate microbubble production, process time could be shorten with the increasing ultrasound power. The obvious ultrasound contrast imaging effects were detected in the dog's heart chamber and liver as well as kidney using only one millilitre agent when diluted. The agent was found safe to the dogs. At the same time, persistent time of the agent was found over 20 min in the dog's body. Conclusion: The new ultrasound contrast agent prepared in the experiment has high microbubble production and concentration, narrow microbubble size distribution ranging in several microns, well stability, little dosage needed in the contrast, well safety to the dogs and long persistent time, obvious contrast imaging effect in the dog's heart chamber, kidney and liver. These experiment data indicate that the new ultrasound contrast agent with three ester surfactants and carboxylic methyl cellulose as its main shell materials can be further developed for clinical purposes.

Experimental system for perfusion imaging and time-intensity processing based on ultrasound contrast agent

Objective： To present a self-developed experimental system for basic studies of blood perfusion imaging and time-intensity evaluating based on ultrasound contrast agent. Methods ： The experimental system performed the image reconstruction and time-intensity processing with radio frequency signals. The system was comprised of ultra-high speed hardware data acquisition interface and low computational cost algorithms. The self-made contrast agent ,blood mimic phantom and capillary phantom model were used to validate the experimental system. Results： The images acquired in blood phantoms with linear-array and curve-array transducers were given. The time-intensity curves corresponding to selected region of interestsequence were demonstrated. It was also shown the time-intensity based decay curves and a decay of ultrasound contrast agent under different ultrasound powers. Conclusion： Several suited from two in vitro phantom models show that the experimental system can be used to f blood perfusion and further clinical studies of microvasculature perfusion.

The combined time-frequency analysis of the acoustic signals backscattered from ultrasonic contrast agents in the evaluation of the blood perfusion

Objective: To analyze the non-periodic, unstable and even chaotic echoes scattered from microbubbles which are extremely sensitive and may easily collapse, fragment or shrink when ultrasound contrast agents are exposed to ultrasound (US) irradiation. Methods: The combined time-frequency analysis was applied to the original signals instead of the traditional Fourier spectral analysis technique. Results: The results obtained from simulation as well as experiment showed that the subharmonic, 2nd harmonic and ultra harmonic of the microbubbles occurred during the oscillation and varied with time. The dependence on the incident ultrasonic amplitude and microbubble parameters were established. Conclusion: The transient echoes backscattered from the ultrasound agent in the evaluation of the blood perfusion can be analyzed thoroughly by the technique of combined-frequency analysis and the time detail of the frequency contents can be revealed.