Method and apparatus for analysis and sorting of polynucleotides based on size

• US 8,388,822 B2
• Filed: 03/22/2012
• Issued: 03/05/2013
• Est. Priority Date: 09/25/1996
• Status: Expired due to Fees

• Current Assignee: California Institute of Technology
• Original Assignee: California Institute of Technology
• Inventors: Quake, Stephen R., Volksmuth, Wayne D.
• Primary Examiner(s): Noguerola, Alex
• Application Number:
  US13/427,815
• Publication Number:
  US 20130001083A1
• Time in Patent Office: 348 Days
• Field of Search: 204601-605, 204451-455
• US Class Current: 204/451
• CPC Class Codes: B01L 2200/0647 Handling flowable solids, e.g. microscopic beads, cells, particles
• CPC Class Codes: B01L 2200/0652 Sorting or classification of particles or molecules
• CPC Class Codes: B01L 2300/0654 Lenses; Optical fibres
• CPC Class Codes: B01L 2300/0816 Cards, e.g. flat sample carriers usually with flow in two horizontal directions
• CPC Class Codes: B01L 2300/0864 comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
• CPC Class Codes: B01L 2400/0415 electrical forces, e.g. electrokinetic
• CPC Class Codes: B01L 2400/0421 electrophoretic flow
• CPC Class Codes: B01L 2400/0484 Cantilevers
• CPC Class Codes: B01L 2400/0487 fluid pressure, pneumatics
• 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 3/502738 characterised by integrated valves microvalves F16K99/0001
• CPC Class Codes: B01L 3/502761 specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads, for physically stretching molecules investigating characteristics of particles G01N15/00
• CPC Class Codes: C12Q 1/6816 characterised by the detection means C12Q1/6804 takes precedence
• CPC Class Codes: C12Q 2565/629 being a microfluidic device
• CPC Class Codes: G01N 15/149 specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 27/44791 Microapparatus sample containers with integrated microfluidic structures B01L3/5027
• CPC Class Codes: Y10S 366/03 Micromixers: variable geometry from the pathway influences mixing/agitation of non-laminar fluid flow

Abstract

The invention relates to a microfabricated device and methods of using the device for analyzing and sorting polynucleotide molecules by size.

Claims

1. A method for transporting and detecting polynucleotides in a microfluidic device, said method comprising:

a) providing a sample comprising polynucleotide molecules of different sizes, wherein said polynucleotide molecules comprise DNA and/or RNA;

b) providing a microfluidic device comprising an inlet, a first microfluidic channel in fluidic communication with a second microfluidic channel, wherein said first and second microfluidic channels join at a junction, and wherein said first microfluidic channel is in fluidic communication with said inlet;

c) transporting said sample into said inlet port and through said first microfluidic channel to said junction with a hydrostatic pressure differential;

d) at said junction electrophoretically diverting said polynucleotides from said first microfluidic channel into said second microfluidic channel;

e) transporting said polynucleotide through said second microfluidic channel; and

f) detecting said polynucleotide by detecting an optically detectable reporter associated with said polynucleotide.

2. The method of claim 1 wherein said polynucleotides are single-stranded DNA molecules.

3. The method of claim 2 wherein the optically-detectable reporter comprises a fluorescent moiety.

4. The method of claim 1 wherein said polynucleotides are single-stranded RNA molecules.

5. The method of claim 4 wherein the optically-detectable reporter comprises a fluorescent moiety.

6. The method of claim 1 wherein the optically-detectable reporter comprises a fluorescent moiety.

7. The method of claim 1 wherein the hydrostatic pressure results from applying pressure at the inlet end.