Patent ID: 8599376
Filing Date: 2013-12-03
Classification: G01N,G02B

Abstract:
1. A method of imaging a sample with an optical system characterized by a diffraction-limited resolution volume, wherein the sample includes a plurality of phototransformable optical labels (“PTOLs”) distributed in the sample with a density greater than an inverse of the diffraction-limited resolution volume of the optical system, the method comprising: (a) providing activation radiation to the plurality of PTOLs to activate a statistical subset of the PTOLs to a state capable of being excited by excitation radiation to a state that emits radiation at a first wavelength, wherein a density of the PTOLs of the activated subset is less than an inverse of the diffraction-limited resolution volume; (b) providing excitation radiation to the activated subset of PTOLs to excite at least some of the activated PTOLs to the state that emits radiation at the first wavelength; (c) detecting the radiation emitted from the activated and excited PTOLs with the optical system, wherein the detecting includes passing the emitted radiation through an objective lens of the optical system; (d) repeating (a) through (c) one or more times, each time activating a different statistical subset of the plurality of PTOLs; and (e) translating a position, within the sample, of a focal plane of the objective lens and then repeating (d); and (f) determining three-dimensional locations within the sample, with a sub-diffraction-limited accuracy, of at least some of the activated and excited PTOLs based on the radiation emitted from the activated and excited PTOLs that is detected when the focal plane of the objective lens is in the different positions.