Single molecule detection system and methods

• US 9,777,321 B2
• Filed: 09/09/2016
• Issued: 10/03/2017
• Est. Priority Date: 03/15/2010
• Status: Active Grant

• Current Assignee: Industrial Technology Research Institute
• Original Assignee: Industrial Technology Research Institute
• Inventors: Chiou, Chung-Fan, Pu, Ying-Chih, Pan, Chao-Chi, Shih, Chih-Tsung, Li, Ming-Chia, Kuo, Chein-Shiu, Chien, Hung-Chi, Chu, Chang-Sheng
• Primary Examiner(s): Warden, Jill
• Assistant Examiner(s): FISHER, BRITTANY I
• Application Number:
  US15/260,773
• Publication Number:
  US 20170009286A1
• Time in Patent Office: 389 Days
• Field of Search: None
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 2561/113 Real time assay
• CPC Class Codes: C12Q 2563/155 Particles of a defined size, e.g. nanoparticles
• CPC Class Codes: G01N 2021/6439 with indicators, stains, dyes, tags, labels, marks
• 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 21/6452 Individual samples arranged in a regular 2D-array, e.g. multiwell plates
• CPC Class Codes: G01N 21/6454 using an integrated detector array
• CPC Class Codes: G01N 21/648 using evanescent coupling or surface plasmon coupling for the excitation of fluorescence
• CPC Class Codes: G01N 21/7703 using reagent-clad optical fibres or optical waveguides using measurement of total internal reflection or attenuated total reflection G01N21/552; optical fibres or waveguides per se G02B
• CPC Class Codes: G01N 2201/08 Optical fibres; light guides
• CPC Class Codes: G01N 2201/12 Circuits of general importance; Signal processing

Abstract

Embodiments encompass a single-molecule detection system and methods of using the detection system to detect an object. Further, embodiments encompass a detection system comprising a movable light coupler, a waveguide, and a light detector. Embodiments further encompass methods of single-molecule detection, including methods of single-molecule nucleic acid sequencing.

Claims

1. A method of sequencing a nucleic acid, comprising:

providing a detection apparatus comprising

a movable light coupler;

a waveguide comprising;

a core layer, and

a first cladding layer,

wherein at least one adapter site for the movable light coupler is formed in at least the first cladding layer; and

a light detector;

providing at least one nucleic acid molecule;

localizing the at least one nucleic acid molecule individually on the movable light coupler;

localizing at the adapter site the movable light coupler on which the at least one nucleic acid is localized;

generating an evanescent field introduced by a light wave propagating along the core layer;

coupling the evanescent field to the movable light coupler, and thereby forming an induced light field around the surface of the movable light coupler;

performing single molecule sequencing-by-synthesis of the at least one nucleic acid molecule, wherein the single molecule nucleic acid sequencing-by-synthesis leads to production of an emitted light correlated to the identity of at least one base in the nucleic acid;

detecting the emitted light with the detector, resulting in an output signal; and

processing the output signal to determine an identity of at least one base comprised by the nucleic acid.

2. The method of claim 1, wherein the refractive index of the movable light coupler is higher than the refractive index of the first cladding layer, and the refractive index of the core layer is higher than the refractive index of the first cladding lay.