INTEGRATED ANALYTICAL SYSTEM AND METHOD

• US 20140171331A1
• Filed: 12/16/2013
• Published: 06/19/2014
• Est. Priority Date: 02/19/2010
• Status: Active Grant

• Current Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Original Assignee: Pacific Bioscience of California Incorporated (L'Oreal SA)
• Inventors: Turner, Stephen, Helgesen, Scott Edward, McCaffrey, Nathaniel Joseph, Saxena, Ravi
• Granted Patent: US 8,867,038 B2
• Time in Patent Office: Days
• US Class Current: 506/6
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2300/0663 Whole sensors
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/168 Specific optical properties, e.g. reflective coatings
• CPC Class Codes: B01L 3/502707 characterised by the manufacture of the container or its components by shaping or joining plastic parts B29C59/00, B29C65/00; by laminating B32B37/00; manufacture of microstructural devices in general B81C
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B82Y 20/00 Nanooptics, e.g. quantum optics or photonic crystals
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 1/6869 Methods for sequencing
• CPC Class Codes: C12Q 1/6874 involving nucleic acid arrays, e.g. sequencing by hybridisation
• CPC Class Codes: G01N 2021/0346 Capillary cells; Microcells
• CPC Class Codes: G01N 2021/6434 Optrodes
• CPC Class Codes: G01N 2021/6439 with indicators, stains, dyes, tags, labels, marks
• CPC Class Codes: G01N 2021/6441 with two or more labels
• CPC Class Codes: G01N 2021/6463 Optics
• CPC Class Codes: G01N 2021/757 using immobilised reagents
• CPC Class Codes: G01N 2021/7786 Fluorescence
• CPC Class Codes: G01N 21/03 Cuvette constructions
• CPC Class Codes: G01N 21/0303 Optical path conditioning in cuvettes, e.g. windows; adapted optical elements or systems; path modifying or adjustment G01N21/031 - G01N21/15 take precedence
• CPC Class Codes: G01N 21/05 Flow-through cuvettes G01N21/09 takes precedence; handling fluid samples G01N1/10
• CPC Class Codes: G01N 21/253 : for batch operation, i.e. multisample apparatus analytical automats G01N35/00
• CPC Class Codes: G01N 21/64 : Fluorescence; Phosphorescence
• 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 21/6454 : using an integrated detector array
• CPC Class Codes: G01N 21/6456 : Spatial resolved fluorescence measurements; Imaging
• CPC Class Codes: G01N 21/648 : using evanescent coupling or surface plasmon coupling for the excitation of fluorescence
• CPC Class Codes: G01N 21/75 : Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated systems in which material is burnt in a flame or plasma G01N21/72, G01N21/73
• CPC Class Codes: G01N 21/77 : by observing the effect on a chemical indicator
• CPC Class Codes: G01N 2201/067 : Electro-optic, magneto-optic, acousto-optic elements
• CPC Class Codes: G01N 2201/068 : Optics, miscellaneous
• CPC Class Codes: G01N 2201/08 : Optical fibres; light guides
• CPC Class Codes: G01N 33/54373 : involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
• CPC Class Codes: G02B 6/1226 : involving surface plasmon interaction
• CPC Class Codes: Y10T 436/143333 : Saccharide [e.g., DNA, etc.]

Abstract

An analytical assembly within a unified device structure for integration into an analytical system. The analytical assembly is scalable and includes a plurality of analytical devices, each of which includes a reaction cell, an optical sensor, and at least one optical element positioned in optical communication with both the reaction cell and the sensor and which delivers optical signals from the cell to the sensor. Additional elements are optionally integrated into the analytical assembly. Methods for forming and operating the analytical system are also disclosed.

Claims

1. A method for sequencing a plurality of template nucleic acid molecules comprising:

adding a sequencing reaction mixture comprising reagents for carrying out polymerase dependent nucleic acid synthesis, including fluorescently labeled nucleotides, to the fluidic layer of an optode array chip;

wherein the optode array chip comprises a two dimensional array of optode elements, each optode element comprising;

a nanoscale aperture within an aperture layer, the aperture forming a fluid receiving nanoscale well for receiving a fluid including sequencing reagents;

above the aperture layer a fluidic layer in fluidic contact with the nanoscale well;

below the aperture layer a waveguide layer that provides illumination to the nanoscale well, the waveguide layer comprising an array of channel waveguides, each channel waveguide extending across a plurality of optode elements;

below the waveguide layer a transmission layer that transmits light emitted from the fluorescent species in the nanoscale well to a detector layer; and

below the transmission layer the detector layer comprising a detector which receives and detects the emitted light from the nanoscale well and transmitted through the transmission layer;

wherein a plurality of the nanoscale wells have within them a single polymerase-template complex comprising a polymerase enzyme and a template nucleic acid.

providing illumination to waveguides on the chip to illuminate optode array elements;

detecting fluorescent light emitted from the nanoscale wells using the detectors in the detector layer;

transmitting electrical signals from the detectors off of the chip through electrical contacts on the chip, and

processing the signals from the detectors in order to sequence a plurality of template nucleic acid molecules.

2. The method of claim 1 wherein each optode element further comprises a lens or an optical tunnel that preferentially directs optical signals from the nanoscale well to the detector.

3. The method of claim 1 wherein the chip comprises from about 1000 to about one million optode elements.

4. The method of claim 1 wherein the detector layer comprises embedded circuits for processing output data from the detector.

5. The method of claim 4 wherein the embedded circuits filter background noise from the detector.

6. The method of claim 4 further comprising an amplifier for amplifying the output data.

7. The method of claim 4 wherein embedded circuits comprise an analog to digital converter.

8. The method of claim 1 wherein there is little or no free space between the aperture, waveguide and transmission layers.

9. The method of claim 1 wherein the detector layer can be reversibly separated from the portion of the array comprising the aperture layer.

11. The method of claim 1 wherein the processing is carried out using a plurality of optional field programmable gate array (FPGA) blocks and application-specific integrated circuits (ASIC).

12. The method of claim 1 wherein the processing is carried out using a central processing unit (CPU) for processing data and controlling the system.

13. The method of claim 1 wherein the transmission layer comprises a filter, prism, grating, or dispersive optical elements to differentially direct different spectral components to different regions of the detector.

14. The method of claim 1 wherein each optode element has one, two, three, or four pixel elements.

15. The method of claim 1 wherein the illumination provided by the waveguide layer is modulated or interleaved to identify two or more different fluorescently labeled nucleotide.