Hong, Sung Min (2012-08). Microfluidics for Single Molecule Detection and Material Processing. Doctoral Dissertation. Thesis uri icon

abstract

  • In the cancer research, it is important to understand protein dynamics which are involved in cell signaling. Therefore, particular protein detection and analysis of target protein behavior are indispensable for current basic cancer research. However, it usually performed by conventional biochemical approaches, which require long process time and a large amount of samples. We have been developed the new applications based on microfluidics and Raster image Correlation spectroscopy (RICS) techniques. A simple microfluidic 3D hydrodynamic flow focusing device has been developed for quantitative determinations of target protein concentrations. The analyte stream was pinched not only horizontally, but also vertically by two sheath streams by introducing step depth cross junction structure. As a result, a triangular cross-sectional flow profile was formed and the laser was focused on the top of the triangular shaped analyte stream. Through this approach, the target protein concentration was successfully determined in cell lysate samples. The RICS technique has been applied to characterize the dynamics of protein 53 (p53) in living cells before and after the treatment with DNA damaging agents. P53 tagged with Green Fluores-cent Protein (GFP) were incubated with and without DNA damaging agents, cisplatin or eptoposide. Then, the diffusion coefficient of GFP-p53 was determined by RICS and it was significantly reduced after the drug treatment while that of the one without drug treatment was not. It is suggested that the drugs induced the interaction of p53 with either other proteins or DNA. This result demonstrates that RICS is able to detect protein-protein or protein-DNA interactions in living cells and it may be useful for the drug screening. As another application of microfluidics, an integrated microfluidic platform was developed for generating collagen microspheres with encapsulation of viable cells. The platform integrated four automated functions on a microfluidic chip, (1) collagen solution cooling system, (2) cell-in-collagen microdroplet generation, (3) collagen microdroplet polymerization, and (4) incubation and extraction of the microspheres. This platform provided a high throughput and easy way to generate uniform dimensions of collagen microspheres encapsulating viable cells that were able to proliferate for more than 1 week.
  • In the cancer research, it is important to understand protein dynamics which are involved in cell signaling. Therefore, particular protein detection and analysis of target protein behavior are indispensable for current basic cancer research. However, it usually performed by conventional biochemical approaches, which require long process time and a large amount of samples. We have been developed the new applications based on microfluidics and Raster image Correlation spectroscopy (RICS) techniques.

    A simple microfluidic 3D hydrodynamic flow focusing device has been developed for quantitative determinations of target protein concentrations. The analyte stream was pinched not only horizontally, but also vertically by two sheath streams by introducing step depth cross junction structure. As a result, a triangular cross-sectional flow profile was formed and the laser was focused on the top of the triangular shaped analyte stream. Through this approach, the target protein concentration was successfully determined in cell lysate samples.

    The RICS technique has been applied to characterize the dynamics of protein 53 (p53) in living cells before and after the treatment with DNA damaging agents. P53 tagged with Green Fluores-cent Protein (GFP) were incubated with and without DNA damaging agents, cisplatin or eptoposide. Then, the diffusion coefficient of GFP-p53 was determined by RICS and it was significantly reduced after the drug treatment while that of the one without drug treatment was not. It is suggested that the drugs induced the interaction of p53 with either other proteins or DNA. This result demonstrates that RICS is able to detect protein-protein or protein-DNA interactions in living cells and it may be useful for the drug screening.

    As another application of microfluidics, an integrated microfluidic platform was developed for generating collagen microspheres with encapsulation of viable cells. The platform integrated four automated functions on a microfluidic chip, (1) collagen solution cooling system, (2) cell-in-collagen microdroplet generation, (3) collagen microdroplet polymerization, and (4) incubation and extraction of the microspheres. This platform provided a high throughput and easy way to generate uniform dimensions of collagen microspheres encapsulating viable cells that were able to proliferate for more than 1 week.

publication date

  • August 2012