Abstract
We have developed a new time-resolved fluorescence platform which enables us to follow the molecular orientation and dynamics of a lipid monolayer at the air - water interface. This enables us to identify fluorescence probe orientation and dynamic freedom within our membrane model. We have dubbed our technique Dynamic three-Dimensional Fluorescence Microscopy (D3DFM). We demonstrate the novelty and applicability of this device by contrasting the time-resolved fluorescence signal of two different fluorescent probes: NBD-PC and BODIPY bound to a lipid layer (DPPC) and a fatty acid layer (Stearic Acid). We control the phase behavior of our sample by controlling the pressure, and find that unlike the rotational diffusion, the in-plane wobbling is highly sensitive to the the position along the pressure-area isotherm. We believe this is indicative phase coexistence. Other fluorescence techniques are not sensitive to this form of motion due to the geometric constraints of collinear excitation. Using this probe we are able to characterize local dynamic changes that take place upon lipid phase transition, which may be critical for membrane protein recognition and insertion.
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