3D Localization Microscopy With Ground State Depletion (GSD 3D)

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3D Localization Microscopy With Ground State Depletion (GSD 3D)

Transcript of 3D Localization Microscopy With Ground State Depletion (GSD 3D)

3D Localization Microscopy With Ground State Depletion (GSD 3D)

3D Localization Microscopy With Ground State Depletion (GSD 3D)

Basic Principles of the GSD Super-Resolution Technology for Displaying Highly Resolved Cellular Structures in Three DimensionsThe exact position of a fluorochrome is based on the principle of temporal separation;

This requires switching of fluorochromes between on and off states followed by a sequential readout;

Super-resolution imaging

The crucial step of dSTORM/GSD is to temporarily switch off the majority of fluorophores to allow the precise localization of single fluorophores. Super-resolution as reconstructed z-stackFluorophore in the focal plane: the image appears round; Below the focus: elongated in the vertical direction;Higher than the focus position: elongated in the horizontal direction.

Section localization scheme

Super-resolution as reconstructed z-stackCoordinates in z are assigned to a specific layer in a z-stack based on a calibration of the deformed PSF by the cylinder lens.Resolution around 70 nm can be achieved.

Evolution of resolutionAlexaFluor 488AlexaFluor 532Rhodamine 6GAlexaFluor 647Others.