Atomic force microscopy investigation of biological systems for health and energy [electronic resource]

Unlike scanning electron microscopy and transmission electron microscopy, atomic force microscopy (AFM) can examine biological samples in situ without sample fixation. Besides, sample manipulation such as nano-indentation can yield mechanical properties of biological samples. Additional signal channels such as electric signals can be obtained with a functionalized electrochemical probe. AFM can also overcome the optical wavelength limitation of optical microscope to resolve single molecules. In my research, AFM was used to study biological samples ranging from cell colonies, single cells down to single molecules. The recent technique of transducing key transcription factors into unipotent cells (fibroblasts) to generate pluripotent stem cells (induced pluripotent stem cells [iPSCs]) has significantly changed the stem cell field. These cells have great promise for many clinical applications, including that of regenerative medicine. To investigate differences between different cell lines, I looked at cell stiffness as a possible indicator of cell differentiation-potential differences. I used AFM to determine the mechanical properties of cell colonies including fibroblasts, multipotent human adipose-derived stromal cells (hASCs) and pluripotent cells, including gold standard human embryonic stem cells (hESCs), hASC-iPSCs and fibroblasts-iPSCs. From least to most stiff, the order of cell stiffness was as follows: hASC-iPSC, hESC, fibroblast-iPSC, fibroblasts, and hASC. The change in mechanical properties of the cells in response to reprogramming offers insight into how the cell interacts with its environment and might provide clues to efficiently reprogram cell populations and maintain their pluripotent state. To more efficiently use the solar energy harvested by photosynthetic organisms, we evaluated the feasibility of generating bioelectricity by directly extracting electrons from the photosynthetic electron transport chain before they are used to fix CO2 into sugars and polysaccharides. An open micro-fluidic channel system was fabricated to immobilize individual cells (5 to 10 microns in diameter) in an arrayed fashion for AFM manipulated probe access. From a living algal cell, Chlamydomonas reinhardtii, photosynthetic electrons 1.2 pA at 6000 mA/m^2 were directly extracted without a mediator electron carrier by inserting a nano-electrode into the algal chloroplast and applying an overvoltage. This result may represent an initial step in generating "high efficiency" bioelectricity by directly harvesting high energy photosynthetic electrons. Photosystem II (PSII) is a pigment-protein complex that oxidizes water and reduces plastoquinone during photosynthesis. Non-photochemical quenching (NPQ) is a protection mechanism to dissipate excess light energy as heat in high light conditions to prevent the formation of singlet-oxygen, an extremely damaging reactive species. State transition is the major NPQ process in Chlamydomonas. Mobile light harvesting complex (LHCII) antenna will associate with PSII or move away from PSII in state 1 or state 2, respectively. We examined thylakoid membranes purified from cells in different states with AFM to study the supramolecular reorganization of PSII supercomplexes during state transition. This work will help us understand the mechanism of state transition and shed light on how the photosynthetic apparatus acclimates to environmental changes at the supramolecular level.

Keywords: Mechanical Engineering.
Advisor: Prinz F. B. primary advisor. Thesis advisor
Grossman Arthur (Arthur R.) advisor. Thesis advisor
Pruitt Beth advisor. Thesis advisor