Single molecule tracking

• US 4,793,705 A
• Filed: 10/07/1987
• Issued: 12/27/1988
• Est. Priority Date: 10/07/1987
• Status: Expired due to Term

• Current Assignee: Los Alamos National Security LLC (Government of the United States of America)
• Original Assignee: United States Department of Energy
• Inventors: Shera, E. Brooks
• Primary Examiner(s): McGraw, Vincent P.
• Application Number:
  US07/105,376
• Time in Patent Office: 447 Days
• Field of Search: 356/317, 356/318, 356/417, 250/458.1, 250/459.1, 250/461.1, 250/461.2
• US Class Current: 356/318
• CPC Class Codes: G01N 15/1459 the analysis being performed on a sample stream
• CPC Class Codes: G01N 2015/0038 Investigating nanoparticles
• CPC Class Codes: G01N 2015/0222 from dynamic light scattering, e.g. photon correlation spectroscopy
• CPC Class Codes: G01N 2015/1027 Determining speed or velocity of a particle
• CPC Class Codes: G01N 21/6402 Atomic fluorescence; Laser induced fluorescence
• 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

Abstract

A detection system is provided for identifying individual particles or molecules having characteristic emission in a flow train of the particles in a flow cell. A position sensitive sensor is located adjacent the flow cell in a position effective to detect the emissions from the particles within the flow cell and to assign spatial and temporal coordinates for the detected emissions. A computer is then enabled to predict spatial and temporal coordinates for the particle in the flow train as a function of a first detected emission. Comparison hardware or software then compares subsequent detected spatial and temporal coordinates with the predicted spatial and temporal coordinates to determine whether subsequently detected emissions originate from a particle in the train of particles. In one embodiment, the particles include fluorescent dyes which are excited to fluoresce a spectrum characteristic of the particular particle. Photones are emitted adjacent at least one microchannel plate sensor to enable spatial and temporal coordinates to be assigned. The effect of comparing detected coordinates with predicted coordinates is to define a moving sample volume which effectively precludes the effects of background emissions.

Claims

1. A molecule detection system, comprising:

a flow cell for passing a train of molecules in laminar flow;

laser means for exciting said molecule to emit photons at a selected wavelength;

position sensitive sensor means effective to detect said photon emissions within said flow cell and assign spatial and temporal coordinates for said detected photons;

computer means for predicting spatial and temporal coordinates for a molecule in said laminar flow as a function of a first detected photon; and

comparison means for comparing said detected spatial and temporal coordinates with said predicted spatial and temporal coordinates to determine whether a detected photon originated from an excited molecule in said train of molecules.

2. A detection system according to claim 1, wherein said position sensitive sensor means includes at least one microchannel plate sensor for outputting said spatial coordinates.

3. A detection system according to claim 1, wherein said position sensitive sensor means has a position accuracy effective for said computer means to create from said predicted coordinates a moving sample volume effective to functionally eliminate background signals from consideration.

4. A detection system for identifying individual molecules having characteristic emissions in a flow train of molecules in a flow cell, comprising:

position sensitive sensor means effective to detect said emissions from said molecules within said flow cell and assign spatial and temporal coordinates for said detected emissions;

computer means for predicting spatial and temporal coordinates for a molecule in said flow train as a function of a first detected emission; and

comparison means for comparing said detected spatial and temporal coordinates with said predicted spatial and temporal coordinates to determine whether subsequently detected emissions originate from an excited molecule in said train of molecules.

5. A detection system according to claim 4, wherein said position sensitive sensor means includes at least one microchannel plate sensor for outputting said spatial coordinates.

6. A detection system according to claim 4, wherein said position sensitive sensor means has a position accuracy effective for said computer means to create from said predicted coordinates a moving sample volume effective to functionally eliminate background signals from consideration.

7. A detection method for identifying individual molecules having a characteristic emission in a flow train, comprising:

detecting in position sensitive sensor means molecular emissions within said flow train;

assigning spatial and temporal coordinates for said detected emissions;

predicting spatial and temporal coordinates for a molecule as a function of a first detected emission; and

comparing said detected spatial and temporal coordinates with said predicted spatial and temporal coordinates to determine whether subsequently detected emissions originate from a molecule in said train of molecules.

8. A method according to claim 7, wherein detecting emissions includes the step of focusing products from said emissions on at least one microchannel plate sensor.

9. A method according to claim 7, wherein said predicting spacial and temporal coordinates further defines a moving sample window for molecule detection effective to discriminate background emission events from molecule emission events.

10. A method according to claim 7, wherein said emission products are photons.

11. A method according to claim 10, further including the step of irradiating said molecules with a laser to induce a fluorescence for emitting said photons.

12. A method according to claim 7, further including the step of hydrodynamically focusing said flow train within a flow cell.