Apparatus and methods for sorting particles

• US 7,943,384 B2
• Filed: 06/07/2010
• Issued: 05/17/2011
• Est. Priority Date: 03/28/2003
• Status: Active Grant

• Current Assignee: Inguran LLC
• Original Assignee: Inguran LLC
• Inventors: Westfall, Lon A., Vandre, Gary P., Nayak, Niraj V., Wallace, Jeffrey D., Hatcher, Jeremy T., Durack, Gary
• Primary Examiner(s): Wallenhorst; Maureen M
• Application Number:
  US12/794,921
• Publication Number:
  US 20100248362A1
• Time in Patent Office: 344 Days
• Field of Search: 422/67, 422/68.1, 422/73, 422/82.05, 422/82.08, 436/63, 436/164, 436/165, 436/43, 436/50, 436/55, 436/172, 435/2, 435/287.1, 435/286.1, 435/286.5, 435/366
• US Class Current: 436/63
• 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 flow cytometry system and method for sorting a mixture of particles with a fluid delivery system delivering a fluid containing particles, a flow cytometry apparatus for receiving the fluid, forming the fluid into a stream and using flow cytometry to classify the particles according to certain characteristics; a sorting system for sorting the particles according to the classification and according to a sorting strategy to provide at least one population containing desired particles and; a control responsive to information received from the flow cytometry apparatus for controlling the sorting system to vary its sorting strategy or for controlling the fluid delivery system to vary the rate at which fluid is delivered as a function at least one of purity and quantity of the particles.

Claims

1. A flow cytometry system for sorting a mixture of particles including particles having a characteristic A and particles having a characteristic B comprising:

a fluid delivery system for delivering a fluid containing said particles;

a flow cytometry apparatus for receiving said fluid, forming the fluid into a stream and using flow cytometry to classify the particles according to said characteristics;

a sorting system in communication with the flow cytometry apparatus for sorting the particles according to said classification and according to a sorting strategy to provide at least one population containing desired particles, wherein the desired particles are selected from the group of particles having either characteristic A or characteristic B and;

a control responsive to information received from the flow cytometry apparatus for;

controlling the sorting system to vary its sorting strategy as a function of at least one of the following;

(1) the purity of said at least one population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A particles or characteristic B particles in said at least one so sulation relative to the total suantit of characteristic A sarticles or characteristic B particles in said stream; and

/or

controlling the fluid delivery system to vary the rate at which fluid is delivered as a function of at least one of the following;

(1) the purity of said at least one population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A particles or characteristic B particles in said at least one population relative to the total quantity of characteristic A particles or characteristic B particles in said stream.

2. The system of claim 1 wherein the particles are sperm cells, and wherein characteristic A is indicative of a live X sperm cell.

3. The system of claim 1 wherein the control increases the rate of fluid delivery when said purity is greater than a desired purity and decreases the rate of fluid delivery when said purity is less than said desired purity.

4. A method of using a flow cytometry system to sort a mixture of particles including particles having a characteristic A and particles having a characteristic B, said method comprising:

delivering a fluid containing said particles to a flow cytometer;

forming the fluid into a stream and using the flow cytometer to classify the particles in the stream according to said characteristics;

sorting the particles in the stream according to said classification and according to a sorting strategy thereby to provide at least one population containing desired particles, wherein the desired particles are selected from the group of particles having either characteristic A or characteristic B; and

varying the sorting strategy as a function of at least one of the following;

(1) the purity of said at least one population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A articles or characteristic B particles in said at least one population relative to the total quantity of characteristic A particles or characteristic B particles in said stream; and

/or

varying the rate at which fluid is delivered to said flow cytometer as a function of at least one of the following;

(1) the purity of said at least one population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A particles or characteristic B particles in said at least one population relative to the total quantity of characteristic A particles or characteristic B particles in said stream.

5. The method of claim 4 wherein said particles are sperm cells, and wherein characteristic A is indicative of a live X sperm cell.

6. The method of claim 4 further comprising increasing the rate of fluid delivery when said purity is greater than a desired purity and decreasing the rate of fluid delivery when said purity is less than the desired purity.

7. A system for sorting a mixture of particles including particles having a characteristic A and particles having a characteristic B, said system comprising:

a fluid delivery system for delivering a fluid stream containing said particles;

a flow cytometry apparatus for receiving said stream, forming droplets containing said particles, and sorting said droplets into different populations according to a sorting strategy, said droplets including first droplets each containing one or more particles having characteristic A, second droplets each containing one or more particles having characteristic B and third droplets each containing one or more particles having characteristic A and one or more particles having characteristic B; and

a control responsive to information received from the flow cytometry apparatus for;

controlling the flow cytometry apparatus to vary the sorting strategy as a function of at least one of the following;

(1) the purity of at least one droplet population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A s articles or characteristic B particles in at least one droplet population relative to the total quantity of characteristic A particles or characteristic B particles in said stream; and

/or

controlling the fluid delivery system to vary the rate at which fluid is delivered as a function of at least one of the following;

(1) the purity of at least one droplet population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A particles or characteristic B particles in at least one droplet population relative to the total quantity of characteristic A particles or characteristic B particles in said stream.

8. The system of claim 7 wherein the control controls the fluid delivery system to maintain the rate at which fluid is delivered as substantially constant, and wherein the control varies the sort strategy.

9. The system of claim 8 wherein the control varies the sort strategy in order to vary the percentage of third droplets in one of the droplet populations.

10. The system of claim 7 wherein the control controls the fluid delivery system to vary the rate at which fluid is delivered as a function of the purity of said at least one droplet population.

11. The system of claim 7 wherein the control controls the fluid delivery system to vary the rate at which fluid is delivered as a function of the quantity of characteristic A particles in said at least one droplet population relative to the total quantity of characteristic A particles in said stream.

12. The system of claim 7 wherein said particles are sperm cells, and wherein characteristic A is indicative of a live X sperm cell.

13. The system of claim 7 wherein the ratio of characteristic A particles to characteristic B particles in said mixture is a known ratio, and wherein said control is operable to classify some of the particles as having characteristic A or characteristic B and to vary the fluid delivery rate as a function of the ratio of classified particles to the known ratio.

14. The system of claim 7 wherein the control determines a purity of the sorted particles based on output signals from the flow cytometry apparatus, said system further comprising an operator input to the control for indicating a desired purity, and wherein the control varies the rate at which fluid is delivered so that the purity of said at least one droplet population generally corresponds to the desired purity.

15. A method of sorting a mixture of particles including particles having a characteristic A and particles having a characteristic B, said method comprising:

delivering a fluid stream containing said particles to a flow cytometer forming droplets containing said particles from the fluid stream in the flow cytometer;

sorting said droplets into different populations according to a sorting strategy, said droplets including first droplets each containing one or more particles having characteristic A, second droplets each containing one or more particles having characteristic B and third droplets each containing one or more particles having characteristic A and one or more particles having characteristic B; and

varying the sorting strategy as a function of at least one of the following;

(1) the purity of at least one droplet population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A particles or characteristic B particles in at least one droplet population relative to the total quantity of characteristic A particles or characteristic B particles in said fluid stream; and

/or

varying the rate at which fluid is delivered to the flow cytometer as a function of at least one of the following;

(1) the purity of at least one droplet population with respect to either characteristic A particles or characteristic B particles; and

(2) the quantity of characteristic A particles or characteristic B particles in at least one droplet population relative to the total quantity of characteristic A particles or characteristic B particles in said fluid stream.

16. The method of claim 15 further comprising maintaining the rate at which fluid is delivered as substantially constant, and varying the sorting strategy.

17. The method of claim 16 further comprising varying the sorting strategy in order to vary the percentage of third droplets in one of the droplet populations.

18. The method of claim 15 further comprising varying the rate at which fluid is delivered as a function of the purity of at least one of the droplet populations.

19. The method of claim 15 further comprising varying the rate at which fluid is delivered as a function of the quantity of characteristic A particles in said at least one droplet population relative to the total quantity of characteristic A particles in said fluid stream.

20. The method of claim 15 wherein said particles are sperm cells, and wherein characteristic A is indicative of a live X sperm cell.

21. The method of claim 15 further comprising increasing the rate at which fluid is delivered when the purity of said at least one droplet population is greater than a desired purity and decreasing the rate when the purity of said at least one droplet population is less than the desired purity.

22. A system for sorting X and Y sperm cells, said system comprising:

a variable rate fluid delivery system for delivering a fluid stream containing X and Y sperm cells;

a flow cytometry apparatus for (1) receiving said stream and forming droplets containing said X and Y sperm cells, said droplets comprising first droplets each containing one or more X sperm cells, second droplets each containing one or more Y sperm cells, and third plurality of droplets each containing one or more X sperms cells and one or more Y sperm cells, (2) sorting said first droplets from said second and third droplets, (3) collecting said first droplets to provide at least one population of X sperm cells, and (4) identifying a quantity of X sperm cells in the at least one population; and

a control responsive to instructions received from the flow cytometer apparatus for varying the rate at which the fluid is delivered to the flow cytometer apparatus as a function of the quantity of identified X sperm cells in said at least one population relative to the total number of X cells in said first and third droplets.

23. The system of claim 22 wherein said flow cytometry apparatus is operable to identify the number of X cells collected in said at least one population, and wherein the control varies the rate at which fluid is delivered to the flow cytometer apparatus to maintain the quantity of X cells collected in said at least one population at or above an acceptable quantity relative to the total number of X cells in said first and third droplets.

24. A method of sorting X and Y sperm cells, said method comprising:

delivering a fluid stream containing X and Y sperm cells to a first location and causing said stream to break into droplets at a second location, said droplets comprising first droplets each containing one or more X sperm cells, second droplets each containing one or more Y sperm cells, and third plurality of droplets each containing one or more X sperms cells and one or more Y sperm cells;

sorting said first droplets from said second and third droplets;

collecting said first droplets to provide at least one population of X sperm cells;

identifying a quantity of X sperm cells collected in said at least one population; and

varying the rate at which fluid is delivered to said first location as a function of the quantity of identified X sperm cells collected in said at least one population relative to the total number of X cells in said first and third droplets.

25. The method of claim 24 further comprising varying the rate at which fluid is delivered to said first location to maintain the quantity of X cells collected in said at least one population at or above an acceptable quantity relative to the total number of X cells in said first and third droplets.

26. A system for sorting X and Y sperm cells, said system comprising:

a variable rate fluid delivery system for delivering a fluid stream containing X and Y sperm cells;

a flow cytometer apparatus for (1) receiving said stream and forming droplets containing said X and Y sperm cells, said droplets comprising first droplets each containing one or more X sperm cells, second droplets each containing one or more Y sperm cells, and third plurality of droplets each containing one or more X sperms cells and one or more Y sperm cells, (2) sorting said second and third droplets from said first droplets, (3) collecting said second and third droplets to provide at least one population of Y sperm cells, and (4) identifying a quantity of Y sperm cells in the at least one population; and

a control responsive to instructions received from the flow cytometer apparatus for varying the rate at which the fluid is delivered to the flow cytometer apparatus as a function of the quantity of identified Y sperm cells in said at least one population.

27. The system of claim 26 wherein said control varies the rate at which fluid is delivered to the flow cytometer system to maintain the purity of said at least one population with respect to Y cells at or above a desired purity.

28. A method of sorting X and Y sperm cells, said method comprising:

delivering a fluid stream containing X and Y sperm cells to a first location and causing said stream to break into droplets at a second location, said droplets comprising first droplets each containing one or more X sperm cells, second droplets each containing one or more Y sperm cells, and third plurality of droplets each containing one or more X sperms cells and one or more Y sperm cells;

sorting said second and third droplets from said first droplets;

collecting said second and third droplets to provide at least one population of Y sperm cells;

identifying a quantity of Y sperm cells collected in said at least one population; and

varying the rate at which fluid is delivered to said first location as a function of the quantity of identified Y sperm cells collected in said at least one population.

29. The method of claim 28 further comprising varying the rate at which fluid is delivered to maintain the purity of said at least one population with respect to Y cells at or above a desired purity.