A method and scanning fluorescence microscope for three-dimensional high-resolution imaging of a labeled with fluorophores structure of a sample

For three-dimensional high-resolution imaging of a labeled switchable fluorophores structure in a sample (3), the steps after placing the sample (3) in front of a lens (4) repeatedly executed: directing fluorescence enabling light (34) along the focal plane (5) of the objective (4) to the sample (3) to form along the focal plane (5), a light sheet (19) from the fluorescence enabling light (34), applying of portions of the light sheet (19) with fluorescence prevent light (35) having a zero point within the light sheet (19), wherein the fluorescent prevent light (35) comprises a component (13) which limits the zero point in the direction orthogonal to the focal plane (5), and a further component (14) through the lens (4) onto the sample (3) is directed and limited to the zero in at least one direction of the focal plane (5) of the lens (4); Measuring from the zero of the sample (3) and emitted to the observed one lens (4) fluorescent light (8); Associating the measured fluorescent light (8) to the location of the null point in the sample (3) in a direction orthogonal to the focal plane (5) and in the at least one direction of the focal plane (5) of the lens (4); and displacing the zero point with respect to the sample (3).