Development of Super Sensitive Position Detector and Its Application on the Studies of Plasma Membrane Proteins

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A new-type super-sensitive position detector was developed using a quadrant avalanche photodiode (4APD) as the photo sensor. This position detector was interfaced to a newly-developed reflective polarisation optical microscope with a near-JR (810 nm) external cavity diode Laser as the light source, In this system, temperature and voltage of 4APD can be optimised to achieve maximum S/N ratio.Using colloidal gold particles with a diameter of 10-100 nm as standard samples, 0.5-2 V output signal with>100 S/N was obtained. At this range of signal intensity, bandwidth of position measurement was 0-17 MHz (-3dB).Spatial resolution of the system was examined by moving the standard sample by a PZT stage. At 375x magnification of the microscope, spatial resolution better than 3 nm was achieved. At this time, the spatial resolution is determined by a fluctuation of the power supply, and using more stable power supply, it should be improved to more than 1 nm. Movement within a 200 nm-phi area can be traced.Using this system, lateral movement of plasma membrane proteins were measured. Although membrane proteins which is bound to the cytoskeleton was undetectable in 33 ms video-rate time window, rapid and Browninan diffusion movement restricted within an area of 300-500 nm was observed by more than 1 ms time resolution of the newly developed position detector. Such restricted movement of cytoskeleton-bound proteins suggests flexibility of the cytoskeletal filaments.