Microscopic device and microscopic method for the three-dimensional localization of punctiform objects

摘要

Microscopic device (10) for the three-dimensional localization of point-like objects (14, 16), comprising: two imaging optics (12, 26, 26 ') which each position one and the same point-like object (14, 16) arranged in an object space (18) Image the form of a focus light distribution (40, 40 ', 42, 42') in two separate image spaces (34), two detector units (28, 28 '), each of which is assigned to one of the two imaging optics (12, 26, 26') and in Detection points of a detection surface (27, 27 ') lying in the respective image space (34) detect an evaluable light spot (54, 54') which represents a plane section through the respective focus light distribution (40, 40 ', 42, 42'), and Evaluation unit (60), which sets the detection points of the two detection surfaces (27, 27 ') in pairs in correspondence with one another and by evaluating the two light spots (54, 54') taking into account this detection point correspondence, a lateral xy position of the point-shaped Ob projectes (14, 16) within an object plane lying in the object space (18) and an axial z-position of the point-like object (14, 16) in the direction of an optical axis (O1) perpendicular to the object plane, characterized in that the both imaging optics (12, 26, 26 ') each have an optical means (26, 26') which the respective focus light distribution (40, 40 ', 42, 42') at an angle to one in the respective imaging optics (12, 26, 26 ') aligns the detection axis (36) provided, which is perpendicular to the detection surface (27, 27') of the respective detection unit (26, 26 '), the inclined positions of the two focus light distributions (40, 40) caused by the optical means (26, 26') ', 42, 42') are opposed to each other in such a way that the two light spots (54, 54 ') with a change in the z-position of the point-like object (14, 16) in their respective detection surfaces (27, 27') take into account counter to the detection point correspondence move, and the evaluation unit (60) determines the axial z-position of the point-like object (14, 16) on the basis of the relative position of the two light spots (54, 54 ').