Abstract: An apparatus includes a position-sensitive detector to detect intensities of radiation as a function of position on the detector, and an optical system, characterized by a diffraction-limited resolution volume, adapted for imaging light emitted from activated and excited phototransformable optical labels (“PTOLs”) in a sample onto the position sensitive-detector. A first light source provides activation radiation to the sample to activate a subset of the PTOLs that are distributed in the sample with a density greater than an inverse of the diffraction-limited resolution volume of the optical system. A second light source provides excitation radiation to the sample to excite a portion of the PTOLs in the subset of the PTOLs. A controller controls one both of the activation radiation and the excitation radiation provided to the sample such that a density of PTOLs in the portion of the PTOLs is less than the inverse of the diffraction-limited resolution volume.

Abstract: A method of imaging with an optical system characterized by a diffraction-limited resolution volume is disclosed. In a sample that 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, a first subset of the PTOLs in the sample are activated, and the density of the activated PTOLs in the first subset is less than the inverse of the diffraction-limited resolution volume. A first portion of the PTOLs in the first subset of PTOLs is excited. Radiation emitted from the activated and excited PTOLs in the first portion of PTOLs is detected with the imaging optics, and locations of activated and excited PTOLs in the first portion of PTOLs is determined with a sub-diffraction-limited accuracy based on the detected radiation emitted from the activated and excited PTOLs.

Abstract: An apparatus includes a position-sensitive detector to detect intensities of radiation as a function of position on the detector, and an optical system, characterized by a diffraction-limited resolution volume, adapted for imaging light emitted from activated and excited phototransformable optical labels (“PTOLs”) in a sample onto the position sensitive-detector. A first light source provides activation radiation to the sample to activate a subset of the PTOLs that are distributed in the sample with a density greater than an inverse of the diffraction-limited resolution volume of the optical system. A second light source provides excitation radiation to the sample to excite a portion of the PTOLs in the subset of the PTOLs. A controller controls one both of the activation radiation and the excitation radiation provided to the sample such that a density of PTOLs in the portion of the PTOLs is less than the inverse of the diffraction-limited resolution volume.

Abstract: First activation radiation is provided to a sample that includes fluorescent proteins (“FPs”) to activate a first subset of the FPs in the sample. First excitation radiation is provided to the first subset of FPs in the sample to excite at least some of the activated FPs, and radiation emitted from activated and excited FPs within the first subset of FPs is detecting with imaging optics. The first activation radiation is controlled such that the mean volume per activated FPs in the first subset is greater than or approximately equal to a diffraction-limited resolution volume (“DLRV”) of the imaging optics.

Abstract: Spatially-structured activation radiation having relatively high- and low-intensity regions is provided to a sample that includes phototransformable optical labels (“PTOLs”). A subset of the PTOLs located predominately at relatively high intensity regions of the spatially-structured activation radiation is activated. Spatially-structured excitation radiation is provided to the activated PTOLs, and the excitation radiation is structured so that one or more relatively high intensity regions of the excitation radiation at least partially overlap one or more relatively high intensity regions of the activating radiation to excite PTOLs located at the relatively high intensity excitation radiation regions. Radiation emitted from the activated and excited PTOLs is detected with imaging optics.

Abstract: An optical system includes a substrate adapted for supporting a sample, where the substrate has a refractive index, nsub, larger than a refractive index of the sample, nsample, a source of electromagnetic radiation having wavelength, ?o, and a detector having multiple individual detector elements configured for detecting a signal resulting from an interaction of the electromagnetic radiation with the sample. The system includes dividing optics, directing optics, and control optics. The dividing optics are configured for dividing the electromagnetic radiation from the source into at least three substantially independent excitation beams.

Abstract: First activation radiation is provided to a sample that includes phototransformable optical labels (“PTOLs”) to activate a first subset of the PTOLs in the sample. First excitation radiation is provided to the first subset of PTOLs in the sample to excite at least some of the activated PTOLs, and radiation emitted from activated and excited PTOLs within the first subset of PTOLs is detecting with imaging optics. The first activation radiation is controlled such that the mean volume per activated PTOL in the first subset is greater than or approximately equal to a diffraction-limited resolution volume (“DLRV”) of the imaging optics.

Abstract: A microscope includes a source of electromagnetic radiation, having a wavelength, ?1 and dividing optical elements configured for dividing the radiation from the source into multiple excitation beams. The microscope also includes a detector and directing optical elements, which are configured for directing each excitation beam in unique directions, such that the beams intersect in an excitation region within a sample to create a two-dimensional or three-dimensional interference pattern of multiple excitation maxima within the sample. The detector has individual detector elements, where the detector elements are configured for detecting light resulting from an interaction of an individual excitation maximum and the sample.

Abstract: First activation radiation is provided to a sample that includes phototransformable optical labels (“PTOLs”) to activate a first subset of the PTOLs. Deactivation radiation is provided to the sample to transform activated PTOLs to an unactivated state. The deactivation radiation has a spatially-structured radiation field including intensity minima, such that a second subset of PTOLs located substantially at the intensity minima remain activated, while activated PTOLs exposed to the deactivation radiation outside the minima are transformed into an unactivated form. Excitation radiation is provided to the sample to excite activated PTOLs in the sample located substantially at the intensity minima of the deactivation radiation. Radiation emitted from the activated and excited PTOLs is detected with imaging optics.

Abstract: A method for creating a periodic interference pattern of coherent waves in two or three dimensions, D, includes generating at least D+2 waves, where each wave has substantially the same wavelength, ?, and travels substantially in a unique direction, kn. The waves are directed such that at least a portion of each wave intersects in a common excitation region to create the interface pattern. The directions, kn, of the waves are selected such that the interference pattern within the excitation region forms a Bravais lattice, and at least one wave travels substantially in a direction, ki, such that one or more symmetry operations characteristic of the Bravais lattice map the direction of the wave, kj, onto a direction of a different wave, kj. Tthe directions of the waves, kn, do not all lie on a D-dimensional set of mutually orthogonal axes.

Abstract: An optical system includes a substrate adapted for supporting a sample, where the substrate has a refractive index, nsub, larger than a refractive index of the sample, nsample, a source of electromagnetic radiation having wavelength, ?o, and a detector having multiple individual detector elements configured for detecting a signal resulting from an interaction of the electromagnetic radiation with the sample. The system includes dividing optics, directing optics, and control optics. The dividing optics are configured for dividing the electromagnetic radiation from the source into at least three substantially independent excitation beams.

Abstract: First activation radiation is provided to a sample that includes phototransformable optical labels (“PTOLs”) to activate a first subset of the PTOLs. Deactivation radiation is provided to the sample to transform activated PTOLs to an unactivated state. The deactivation radiation has a spatially-structured radiation field including intensity minima, such that a second subset of PTOLs located substantially at the intensity minima remain activated, while activated PTOLs exposed to the deactivation radiation outside the minima are transformed into an unactivated form. Excitation radiation is provided to the sample to excite activated PTOLs in the sample located substantially at the intensity minima of the deactivation radiation. Radiation emitted from the activated and excited PTOLs is detected with imaging optics.