Apparatus and methods for optical analysis of molecules

• US 7,170,050 B2
• Filed: 09/17/2004
• Issued: 01/30/2007
• Est. Priority Date: 09/17/2004
• Status: Active Grant

• Current Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Original Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Inventors: Korlach, Jonas, Turner, Stephen
• Primary Examiner(s): Wells; Nikita
• Application Number:
  US10/944,106
• Publication Number:
  US 20060060766A1
• Time in Patent Office: 865 Days
• Field of Search: None
• US Class Current: 250/251
• CPC Class Codes: C12Q 1/6806 Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: G01N 2021/6439 with indicators, stains, dyes, tags, labels, marks
• CPC Class Codes: G01N 2021/6463 Optics
• CPC Class Codes: G01N 2021/7786 Fluorescence
• CPC Class Codes: G01N 21/00 Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light G01N3/00 - G01N19/00 take precedence
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/645 Specially adapted constructive features of fluorimeters
• CPC Class Codes: G01N 21/6452 Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• CPC Class Codes: G01N 21/648 using evanescent coupling or surface plasmon coupling for the excitation of fluorescence
• CPC Class Codes: G01N 21/77 by observing the effect on a chemical indicator
• CPC Class Codes: G01N 2201/06113 Coherent sources; lasers

Abstract

The present invention relates to optical confinements, methods of preparing and methods of using them for analyzing molecules and/or monitoring chemical reactions. The apparatus and methods embodied in the present invention are particularly useful for high-throughout and low-cost single-molecular analysis.

Claims

1. A substrate comprising:

an array of a plurality of optical confinements, wherein each of the plurality of optical confinements comprises a core disposed within a cladding layer that is configured to preclude propagation of electromagnetic radiation through the core, and wherein each core is structured to receive reactants; and

wherein the plurality of optical confinements having a density of greater than 4×

10⁴ confinements per mm² are aligned upon the substrate in at least a first row or a first column, wherein said first row or said first column is aligned along at least a first axis of the substrate.

2. The substrate of claim 1, wherein the plurality of optical confinements are present on the substrate at a density of at least 8×

10⁴ confinements per mm².

3. The substrate of claim 1, wherein the plurality of optical confinements are present on the substrate at a density of at least 1.2×

10⁵ confinements per mm².

4. The substrate of claim 1, wherein the array of optical confinements comprises at least 2 rows of optical confinements.

5. The substrate of claim 1, wherein the array of optical confinements comprises at least 10 rows of optical confinements.

6. The substrate of claim 1, wherein the array of optical confinements comprises at least 100 rows of optical confinements.

7. The substrate of claim 4, wherein the at least two rows of optical confinements are further aligned along at least a second axis.

8. The substrate of claim 4, wherein the at least two rows of optical confinements are further aligned in columns.

9. The substrate of claim 4, wherein the at least two rows of optical confinements are further aligned in a hexagonal array.

10. The substrate of claim 1, wherein the optical confinements each comprise a core disposed through a cladding layer that is disposed upon a supporting substrate.

11. The substrate of claim 10, wherein the core comprises a hole disposed through the cladding layer.

12. The substrate of claim 10, wherein the cladding layer comprises a metal layer.

13. The substrate of claim 10, wherein the cladding layer comprises one or more material selected from aluminum, antimony, arsenic, beryllium, bismuth, boron, cadmium, calcium, carbon, cerium, chromium, cobalt, copper, gallium, indium, iron, lead, lithium, magnesium, manganese, mercury, molybdenum, nickel, niobium, phosphorus, silicon, silicate, silicon nitride, gallium phosphide, gallium arsenide, vanadium, and zinc.

14. The substrate of claim 12, wherein the metal layer comprises one or more metal selected from aluminum and chromium.

15. An optical system, comprising:

a substrate comprising;

an array of a plurality of optical confinements, wherein each of the plurality of optical confinements comprises a core disposed within a cladding layer that is configured to preclude propagation of electromagnetic radiation through the core, and wherein each core is structured to receive reactants; and

wherein the plurality of optical confinements are aligned upon the substrate in at least a first row or column that is aligned along at least a first axis of the substrate;

an excitation light source positioned to direct excitation light at the array of optical confinements; and

a charge coupled device (CCD) positioned to detect emitted signals from the array of optical confinements.

16. The optical system of claim 15, wherein the charge coupled device is selected from an electron multiplying charge coupled device and an intensified charge coupled device (ICCD).

17. The optical system of claim 15, further comprising an optical transmission element for directing excitation light at the array of optical confinements and directing emitted light from the array of optical confinements to the CCD.

18. The optical system of claim 15, wherein the array of confinements and the CCD are aligned so that an optical signal from each confinement is directed to a separate photon detector on the CCD.

19. The optical system of claim 17, wherein the CCD is an electron multiplying CCD (EMCCD).

20. The optical system of claim 17, wherein the CCD is an intensified CCD (ICCD).