High-resolution Imaging Study Of Organic/Nanoparticle-coated Micro/Nano Bubbles

Ultrasound imaging（USI）diagnostic technology has become one of the most widely used medical diagnostic technologies because of its advantages of safety,wide application,real-time,repeatable inspection,strong ability to identify soft tissues and high flexibility.As a commonly used contrast agent for ultrasound imaging,organic-coated bubbles,especially micro/nano bubble systems,show very good imaging contrast in USI.In recent years,researchers has developed a combination of microbubble USI contrast agent and superparamagnetic iron oxide nanoparticles（SPIONs）which is magnetic resonance imaging（MRI）contrast agent,a dual-mode（USI and MRI）contrast agent system-organic/nanoparticle-coated micro/nano bubbles studied in this thesis,to obtain very good imaging effects of USI and MRI.In addition,this bimodal contrast agent can be used as a drug carrier in medical diagnosis and treatment.However,such a dual-mode contrast agent needs to meet a condition that entering the tissue must maintain a certain stability to effectively perform its role.How to evaluate its stability is a very important topic.In addition,why bubbles can achieve very good imaging contrast when they reach sub/micron scale?What is the internal structure?However,because their sizes are only few microns or less,it is difficult to detect them.With the development of high-resolution imaging technology,such as atomic force microscopy（AFM）and synchrotron radiation soft X-ray spectroscopy microscopy,it is expected to solve the aforementioned scientific problems.Therefore,this thesis combined with transmission electron microscopy（TEM）,using advanced AFM（PF-QNM mode）to image air micro/nano bubbles coated with PLGA/Fe₃O₄nanoparticles,and analyzed single micro/nano bubbles,evaluated its stability;using advanced synchrotron radiation technology-scanning transmission soft X-ray microscopy（STXM）to study the shell composition and internal chemical information of organic/nanoparticle-coated micro/nano bubbles and initially explored the process of magnetic nano bubbles entering the cell.These studies will provide important guidance for its applications in medical imaging and targeted drug delivery.The main research progress includes two parts.In the first part,we used AFM PF-QNM mode to study the mechanical properties of single PLGA/Fe₃O₄ nanoparticle-coated sub-micron bubbles.A series of force curves were obtained by scanning the sub-micron bubbles coated with PLGA/Fe₃O₄ nanoparticles under different loaded forces.The elastic deformation,inelastic deformation and rupture deformation processes of the bubble were analyzed from their force curves.The elastic modulus and stiffness information of single bubble were calculated under small deformation area and scanning loaded force.The results showed that the elastic modulus of organic/nanoparticle-coated sub-micron bubbles is 4.6±1.2 GPa,which is independent of their sizes,and is between the elastic modulus of inorganic hollow spheres and pure organic polymer microspheres.Its stiffness is 15.0±3.1 N/m,which is obviously higher than the stiffness of the interfacial nano-bubbles,and smaller than the stiffness of inorganic hollow spheres.These results showed that this coating combination strengthens the mechanical properties of the bubbles and makes them have better stability.In addition,TEM imaging also found that most of the coated micro/nano bubbles maintain a very good shape,which also shows that they have good stability.These experimental results would provide the important experimental basis for evaluating the stability of the bimodal contrast agent and drug delivery when they enter living body.In the second part,using STXM we explored the shell composition and internal chemical information of organic/nanoparticle-coated micro/nano bubbles.In the experiment,we first imaged the organic/Fe₃O₄ nanoparticle-coated oxygen micro/nano bubbles.Different elements have different absorption peaks for X-ray.We did a dual-energy contrast scan of the bubbles at the energy of the X-ray absorption peak and before weak absorption of iron L edge to obtain the distribution of iron elements on the bubbles.The distribution of the iron element is also the distribution of Fe₃O₄ nanoparticles,indicating that organic/Fe₃O₄nanoparticle-coated micro/nano bubbles’shell is composed with organic matter and nanoparticles.Then the stack scanning analysis of the bubbles was performed on the K edge of oxygen absorption,and the chemical information inside the bubbles was obtained,proving the existence of oxygen inside the organic/Fe₃O₄ nanoparticle-coated micro/nano bubbles.Also,the experimental results showed that under continuous X-ray transmission scanning,the organic polymer of the bubble shell is significantly damaged by radiation.This indicated that the coated bubbles are unstable under the irradiation of soft X-rays when their size exceeds a certain size（such as larger than 2microns）.This study provides the first detailed structural and chemical information on the interior of this bimodal contrast agent.