Advanced volumetric microscopy techniques for dynamic studies of anucleate cells
Date
2018
Authors
He, Xuefei
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Abstract
Optical imaging is a cornerstone in biological and biomedical sciences. It is imperative to not only obtain high-resolution images but also provide real-time quantification of biological measurement using automated non-invasive imaging solutions. In recent years, the advent of volumetric microscopes, especially laser scanning microscopy (LSM) and quantitative phase microscopy (QPM), has opened up new powerful imaging tools for biological studies.
One of the primary biological activities that require volumetric imaging is blood dynamics. Blood is an essential component in the circulatory system, which transports nutrients, oxygen, and waste for the entire body. Anucleate blood cells, red blood cells (RBCs) and platelets, are majority contributors in homeostasis, the state of steady internal conditions, and hemostasis, the process of bleeding control. Any morphological anomalies of RBCs and platelets could result in mortality. For examples, poor deformability of RBCs influenced by blood-borne diseases (e.g. malaria, sickle) can result in poor delivery of nutrients and oxygen to major visceral organs. And excessive or insufficient blood coagulation due to kinds of platelet disorders can lead to thrombosis or abnormal bleeding.
This thesis presents the development of volumetric imaging systems and tailored microfluidics platform for the study of diseased anucleate blood cells. Using a custom-built automated QPM system, quantitative deformability measurement of parasites infected RBCs was conducted using an adhesion assay. Furthermore, real-time QPM imaging of platelets aggregation was achieved for studying thrombus dynamic. Then, a hybrid image system, Holographic inverted Single Plane Illumination Microscopy (H+iSPIM), was developed to retrieve both structural and molecular information, providing multidimensional images at the same time.
The first chapter introduces the aims and the structure of the thesis. The second, third and fourth chapters presents background of the techniques used in the thesis; volumetric microscopy techniques, biomechanics of anucleate blood cells, and microfluidic respectively. Chapter five elaborates the hardware and software development for an automated DHM system. This chapter is meant to extend the usage of DHM by automated image processing and data analysis. The next following two chapters (six and seven) describe the deformability studies of infected RBC and dynamics of platelet-platelet aggregate. Chapter eight proposes a hybrid imaging system, H+iSPIM, for multidimensional studies of cells in microfluidic systems. Conclusions, future works, and limitations of the developed system are discussed in the final chapter.
In summary, this thesis presents the pursuit of versatile imaging systems for non-invasive imaging quantification of dynamic cellular studies using microfluidic devices for blood-related diseases.
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volumetric microscopy techniques, digital holographic microscope, microfluidics, light sheet microscope
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Thesis (PhD)
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