In situ resonance energy transfer microscopy: monitoring membrane fusion in living cells.

摘要: Publisher Summary The resolution limit of fluorescence light microscopy (about 400 nm) can be exceeded indirectly by using resonance energy transfer (RET) microscopy. This technique is able to visualize the spatial location two different, fluorescently labeled membrane probes and determine if they are in same membrane, or physically adjacent but separate bilayers (10 nm apart. Resonance was developed study temporal distribution fluorescent model membranes living cells. Particular attention paid mechanics visualizing definitively colocalization both bilayer. microscope configuration used ATP-dependent liposome fusion with Golgi apparatus permeabilized, cultured skin fibroblasts. However, combining RET low light-level detector technology digital image analysis will facilitate acquisition kinetic studies. use two-stage three-stage charge coupled cameras allow illumination levels reduce photo bleaching undetectable levels. Microprocessor-controlled imaging quantitatively intensity at discrete locations microscopic field interest calculate wavelength ratios