Ultrahigh resolution multi-color colocalization of single fluorescent probes

a fluorescent probe and multi-color technology, applied in the field of ultra-high resolution multi-color colocalization of single fluorescent probes, can solve the problems of requiring subsequent signal processing, each method has particular limitations, and the deconvolution algorithm does not have unique solutions, so as to improve the spatial resolution of optical microscopy, the effect of demanding operation

a fluorescent probe and multi-color technology, applied in the field of ultra-high resolution multi-color colocalization of single fluorescent probes, can solve the problems of requiring subsequent signal processing, each method has particular limitations, and the deconvolution algorithm does not have unique solutions, so as to improve the spatial resolution of optical microscopy, the effect of demanding operation

US20050123979A1Inactive Publication Date: 2005-06-09WEISS SHIMON +2
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  • Ultrahigh resolution multi-color colocalization of single fluorescent probes
  • Ultrahigh resolution multi-color colocalization of single fluorescent probes
  • Ultrahigh resolution multi-color colocalization of single fluorescent probes

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Embodiment Construction

[0064] The instant invention contemplates a method for the colocalization of two or more species of interest in a sample comprising:

[0065] a) exciting the species of interest with a single wavelength of light thereby causing the two or more species of interest to emit light of a distinctive emission characteristic,

[0066] b) separating the distinctive emission characteristics of the two or more species of interest,

[0067] c) detecting the emitted light from the two or more species of interest,

[0068] d) moving the sample a predetermined distance,

[0069] e) repeating steps a) through d) a predetermined number of times thereby creating a multitude of representations of the excitation point spread function (PSF),

[0070] f) determine the geometric center of the representations of the excitation PSF for at least two of the species of interest,

[0071] g) computing the distance between the geometric centers of the representations of the excitation PSF for the species of interest.

[0072] T...

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Abstract

A novel optical ruler based on ultrahigh-resolution colocalization of single fluorescent probes is described. Two unique families of fluorophores are used, namely energy-transfer fluorescent beads and semiconductor nanocrystal (NC) quantum dots, that can be excited by a single laser wavelength but emit at different wavelengths. A novel multicolor sample-scanning confocal microscope was constructed which allows one to image each fluorescent light emitter, free of chromatic aberrations, by scanning the sample with nanometer scale steps using a piezo-scanner. The resulting spots are accurately localized by fitting them to the known shape of the excitation point-spread-function of the microscope.

Description

CROSS REFERENCE TO RELATED APPLICATIONS [0001] This application claims priority to U.S. Ser. No. 60 / 228,107, filed Aug. 24, 2000, the contents of which are incorporated herein in their entirety.STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT [0002] This invention was made with government support under Grant (Contract) No. DE-AC03-76F00098 awarded by The United States Department of Energy. The government has certain rights to this invention.BACKGROUND OF THE INVENTION [0003] Following the completion of the human genome project, the cataloging of all gene sequences and the acquisition of high-resolution structures of proteins and RNAs, future biological investigations will focus on how the fundamental cellular building blocks interact with each other. Another important issue will be to determine their precise locations in space and time in an attempt to decode and lay out the cell machinery and circuitry. Indeed, many vital functions of the cell are performed by highly...

Claims

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Application Information

Patent Timeline
09 Jun 2005
Publication
US20050123979A1
IPC
G02B21/00
CPC
B82Y15/00; G02B21/008; G02B21/0076; G02B21/0024
Inventors
WEISS, SHIMON; MICHALET, XAVIER