Flow cytometer method and apparatus

• US 8,709,825 B2
• Filed: 03/01/2013
• Issued: 04/29/2014
• Est. Priority Date: 03/28/2003
• Status: Active Grant

• Current Assignee: Inguran LLC
• Original Assignee: Inguran LLC
• Inventors: Durack, Gary, Wallace, Jeffrey D., Vandre, Gary P., Westfall, Lon A., Hatcher, Jeremy T., Nayak, Niraj V.
• Primary Examiner(s): Wallenhorst, Maureen
• Application Number:
  US13/782,459
• Publication Number:
  US 20130224787A1
• Time in Patent Office: 424 Days
• Field of Search: 422/67, 422/68.1, 422/73, 422/82.05, 422/501, 422/509, 422/515, 436/63, 436/164, 436/165, 436/43, 436/50, 436/55, 436/180, 435/2, 435/287.1, 435/286.4, 435/288.7, 435/366, 435/29
• US Class Current: 436/180
• CPC Class Codes: A01N 1/0284 Temperature processes, i.e. using a designated change in temperature over time
• CPC Class Codes: C12N 5/06 Animal cells or tissues; Human cells or tissues preservation of living cells or tissues A01N1/02
• CPC Class Codes: C12N 5/061 Sperm cells, spermatogonia
• CPC Class Codes: C12N 5/0612 sorting of gametes, e.g. according to sex or motility
• CPC Class Codes: C12Q 1/02 involving viable microorganisms
• CPC Class Codes: C12Q 3/00 Condition responsive control processes apparatus therefor C12M1/36; controlling or regulating in general G05
• CPC Class Codes: G01N 15/01 specially adapted for biological cells, e.g. blood cells investigating sedimentation of particle suspensions in blood G01N15/05
• CPC Class Codes: G01N 15/1404 Handling flow, e.g. hydrodynamic focusing
• CPC Class Codes: G01N 15/1459 the analysis being performed on a sample stream
• CPC Class Codes: G01N 15/1468 with spatial resolution of the texture or inner structure of the particle
• CPC Class Codes: G01N 15/147 the analysis being performed on a sample stream
• CPC Class Codes: G01N 15/149 specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 2015/1006 for cytology
• CPC Class Codes: G01N 2015/1406 Control of droplet point
• CPC Class Codes: G01N 2015/1415 Control of particle position
• CPC Class Codes: G01N 2021/6439 with indicators, stains, dyes, tags, labels, marks
• CPC Class Codes: G01N 21/63 optically excited
• 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 33/48 Biological material, e.g. blood, urine G01N33/02, G01N33/26, G01N33/44, G01N33/46 take precedence; Haemocytometers counting blood corpuscules distributed over a surface by scanning the surface G06M11/02
• CPC Class Codes: G01N 33/5005 involving human or animal cells immunoassay G01N33/56966; immunoassays of protozoa G01N33/56905; protozoa in screening assays C12Q1/025
• CPC Class Codes: Y10T 436/10 : Composition for standardization, calibration, simulation, stabilization, preparation or preservation; processes of use in preparation for chemical testing
• CPC Class Codes: Y10T 436/115831 : Condition or time responsive
• CPC Class Codes: Y10T 436/12 : Condition responsive control
• CPC Class Codes: Y10T 436/25 : including sample preparation
• CPC Class Codes: Y10T 436/2525 : Stabilizing or preserving
• CPC Class Codes: Y10T 436/25375 : Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]
• CPC Class Codes: Y10T 436/255 : including use of a solid sorbent, semipermeable membrane, or liquid extraction
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

A nozzle for use in a flow cytometer and a method of orienting particles with a baffle disposed within a nozzle. The baffle may be a baffle plate positioned in a fluid flow path to deflect a fluid stream and to orient particles in a desired orientation as they exit the nozzle.

Claims

1. A nozzle for use in flow cytometry apparatus for analyzing particles in a fluid stream, said fluid stream comprising a sheath stream surrounding a core stream containing said particles, said nozzle comprising:

a nozzle having an interior surface defining a flow path for said fluid stream, and an orifice for exit of the fluid stream from the nozzle;

a sample injection needle disposed within the nozzle; and

a baffle plate located downstream of the sample injection needle in the nozzle positioned in said flow path upstream from said orifice for deflecting the fluid stream as it moves along said flow path, the baffle plate and interior surface of the nozzle being configured to orient the particles in a desired orientation as they exit the orifice.

2. The nozzle of claim 1, wherein said nozzle has a first cross sectional flow area upstream of said baffle plate and a second cross sectional flow area different from said first cross sectional flow area at the baffle plate.

3. The nozzle of claim 2, wherein said second cross sectional flow area is smaller than said first cross sectional flow area.

4. The nozzle of claim 2, wherein said first and second cross sectional flow areas have different shapes.

5. The nozzle of claim 4, wherein said second cross sectional flow area is generally semi-cylindrical.

6. The nozzle of claim 1, wherein said interior surface of the nozzle is shaped to define at least two axially tapered regions downstream from said baffle plate for progressively accelerating the speed of the fluid stream in a downstream direction toward the nozzle orifice.

7. The nozzle of claim 6, wherein said baffle plate is configured for deflecting said core stream toward a portion of said interior surface of the nozzle defining at least one of said at least two axially tapered regions.

8. The nozzle of claim 7, wherein said at least two axially tapered regions have generally elliptical cross sectional shapes oriented in different directions for applying torsional forces to said fluid stream that tend to bring the particles into said desired orientation.

9. The nozzle of claim 8, wherein said generally elliptical cross sectional shape of one of said at least two regions is oriented at about 90 degrees relative to the generally elliptical cross sectional shape of the other of said at least two regions.

10. The nozzle of claim 1, wherein the nozzle has an exterior surface with a non-reflective coating thereon.

11. The nozzle of claim 1, further comprising a holder for holding said baffle plate in fixed position in said nozzle.

12. The nozzle of claim 11, wherein said baffle plate has a first leg extending generally transversely relative to said fluid stream and a second leg extending generally in the direction of said fluid stream.

13. The nozzle of claim 12, wherein said interior surface of the nozzle has a region of generally elliptical cross section downstream from said baffle plate, and wherein said first and second legs of the baffle plate intersect along a line which is substantially parallel with a major axis of said generally elliptical cross section.

14. The nozzle of claim 12, wherein said interior surface of the nozzle has a region of generally elliptical cross section downstream from said baffle plate, and wherein said first and second legs of the baffle plate intersect along a line which is substantially perpendicular to a major axis of said generally elliptical cross section.

15. The nozzle of claim 11, wherein said holder comprises a generally cylindrical shell holding said baffle plate and positioned in said nozzle.

16. The nozzle of claim 11, wherein the baffle plate has a semi-circular shape or has semi-elliptical shape.

17. The nozzle of claim 11, wherein the interior surface of the nozzle is shaped to define a shoulder and the holder comprises an o-ring seal capable of pressing the baffle plate against the shoulder to hold the baffle plate in a desired position.

18. The method of claim 11, wherein the baffle holder holds the baffle plate in a position in which the baffle plate intersects a longitudinal axis of the nozzle.

19. A method of orienting particles in a flow cytometry apparatus, said method comprising:

forming a fluid stream by introducing a core stream of sample fluid containing particles into a sheath fluid so that the sheath fluid surrounds the core stream;

flowing the fluid stream having a core stream and sheath fluid through a nozzle having an interior surface defining a flow path that generally tapers from an upstream portion of the nozzle to an orifice at the downstream end of the nozzle; and

deflecting the core stream with a baffle plate as it flows through the nozzle to subject the particles in the core stream to hydrodynamic forces that tend to cause the particles to assume a desired orientation.

20. The method of claim 19, wherein the nozzle has a longitudinal axis and the step of deflecting the core stream comprises deflecting the core stream away from the longitudinal axis toward said interior surface with said baffle plate in the nozzle positioned in said flow path.

21. The method of claim 19, wherein the nozzle has a longitudinal axis and the step of deflecting the path of the core stream comprises deflecting the core stream from a path that generally coincides with the longitudinal axis to a deflected path, at least a portion of the deflected path being offset from the longitudinal axis of the nozzle.

22. The method of claim 19, wherein the nozzle has a longitudinal axis, the method further comprising introducing the core stream into the sheath fluid at a location that is offset from the longitudinal axis.

23. The method of claim 19, wherein the nozzle has an interior surface that is shaped to define at least one torsional zone comprising a tapered region having a generally elliptical cross section for subjecting the particles to hydrodynamic orienting forces.

24. The method of claim 23, wherein said interior surface is shaped to define multiple torsional zones.

25. The method of claim 24, wherein each of said multiple torsional zones comprises a tapered region of the nozzle having a generally elliptical cross section, wherein the generally elliptical cross section for a first of the multiple torsional zones is oriented in a different direction than the generally elliptical cross section for a second of the multiple torsional zones.