Design and development of a microfluidic device to monitor iron binding dynamics in iron transport proteins

Official URL: http://risc01.sabanciuniv.edu/record=b1620291 (Table of Contents)

Iron binding mechanisms of proteins are riveting studies because of the great importance of iron molecules for the metabolism of humans and other organisms. Organisms have developed different mechanisms to catch iron from the environment. Although it is known that this mechanism is rapid and efficient, there is lack of kinetic rate data under different environmental conditions to explain the details of the mechanism. In this dissertation, a microfluidic device was designed and developed to measure iron binding constants in ferric binding proteins of humans (Transferrin) and bacteria (Haemophilus Influenzae Ferric Binding Protein (FBP)), to assess the effect of different environmental conditions on the kinetics of iron – protein association. This study aims to contribute the field by providing a cheap and efficient experimental setup that measures reaction rates of iron binding proteins. Firstly, a microfluidic chip housing an effective mixing component was designed and fabricated by using PMMA and PDMS as a material. Both designs were tested by using bromocresol green – acetic acid reactions, where the color change from blue to green and from green to yellow can be observed by lowering the pH. The reaction was monitored with high resolution camera. Color changing property of the reaction was used to illustrate total mass transfer in the mixing chamber to determine dead time. Using ANSYS Fluent software these geometries were modified and improved designs were suggested. Secondly, because the bacterial FBP is not commercially available, it was expressed and purified by using recombinant DNA technology for monitoring iron binding dynamics in the microfluidic device as future work.