Quantitative Phase Imaging For Cellular Biology PDF Download

Quantitative phase imaging using Digital Holographic Microscopy (DHM) is emergine as a label-free and wide-field method of characterizing cells with high spatio-temporal resolution. In parallel, silicon based michromechanical and electronic devices are allowing both manipulation (e.g. electrical stimulation, mechanical actuation) as well as characterization (electrical and mechanical) of micro and nano-scopic samples. THis has revolutionized development of lab-on-a-chip devices for high throughput analysis of cells and molecules for diagnosis of disease and screening of drug-effects. However, very little progress has been made in optical (e.g. fluorescence, Raman etc) characterization of samples on these silicon-based devices. Especially, wide-field high-resolution optical imaging and characterization of samples under silicon environment has not been possible owing to the opacity of silicon to visible light. This thesis reports high resolution near-infrared quantitative phase imaging of cells through silicon, in isotonic as well as hypotonic environment using DHM. Further, several microscopic (AFM, laser manipulation) methods are being developed for characterization of mechanical properties (e.g. elasticity) of cells so as to determine changes during physiological stress. In particular, optical tweezers are used for transverse-stretching cells by actuating anchored-beads as handles and imaging using phase-contrast microscopy. While this method is constantly gaining more attention due to non-contact nature of actuation, it is very time consuming and has low working distance. The thesis describes development of a weakly-focused laser beam for axial-stretching. Application of DHM allowed cell imaging with nm-resolution when stretched axially. Development of an empirical formula for force exerted by defocused light beam on a cell surface led to measurement of elastic property of cell. In addition to this, the thesis aimed at evaluating changes in elastic properties of cell under over-expression of certain proteins (HOX-B9), which is believed to be involved in tumorigenesis. Significant reduction in elastic property of cells over-expressing HOXB9 was found as compared to the control cells. Thus, the thesis paves the way for development of a method for optical manipulation and imaging of cells for characterization of their elastic properties in different physiological states, and probe nanoscale interatctions with different physio-chemical agents in a non-invasis and label-free manner.