METHODS AND SYSTEMS FOR ASSESSING SAMPLE BEHAVIOR IN A FLOW CYTOMETER

• US 20140216128A1
• Filed: 01/31/2014
• Published: 08/07/2014
• Est. Priority Date: 02/01/2013
• Status: Active Grant

• Current Assignee: Becton, Dickinson & Co
• Original Assignee: Becton, Dickinson & Co
• Inventors: Iyer, Sujata, Trotter, Joseph
• Granted Patent: US 10,190,963 B2
• Time in Patent Office: Days
• US Class Current: 73/19.01
• 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 15/149 specially adapted for sorting particles, e.g. by their size or optical properties
• CPC Class Codes: G01N 2015/1006 for cytology

Abstract

Methods and systems are disclosed for generating an entrainment factor in a flow cytometry sample. The methods comprise flowing a sample with a series of particles through the flow cytometer, detecting events and calculating an expected frequency of those events based on a distribution, such as a Poisson distribution, and measuring an observed frequency of particle events. An entrainment factor may be generated from a ratio of observed event frequency to expected event frequency. Further adjustment to the flow cytometer maybe performed based on the indicated entrainment factor such as adjusted sorting bias.

Claims

1. A method for generating an entrainment factor for a flow cytometer sample, the method comprising:

flowing the sample comprising a series of particles through the flow cytometer wherein each particle is associated with a signal;

detecting a series of events with a detection system in a first time interval from a stream formed from the flow cytometer;

calculating an expected frequency of events in the first time interval based on a characterizing distribution;

measuring an observed frequency of events within a second time interval, wherein the second time interval is within the first time interval; and

generating the entrainment factor based at least in part on a ratio of the observed frequency to the expected frequency.

2. The method of claim 1, wherein the characterizing distribution includes a Poisson distribution.

3. The method of claim 1, further comprising generating at least one of an estimated yield or an estimated purity based on the entrainment factor and a sorting logic.

10. The method of claim 1, wherein the entrainment factor is generated while the sample is flowing in the flow cytometer.

12. The method of claim 1, wherein the entrainment factor is one of a plurality of entrainment factors calculated during successive time intervals for a flow cytometer sample.

13. The method of claim 1, wherein the sample comprises peripheral blood cells wherein the peripheral blood cells as are comprised of a first and second component labeled with a first and second label.

14. The method of claim 13, wherein the entrainment factor is generated for the first component and another entrainment factor is generated for the second component.

4. A method for generating sample behavior information for a flow cytometer sample, the method comprising:

flowing the sample comprising a series of particles through the flow cytometer wherein each particle is associated with an event and the particles are distributed in a fluidic stream;

detecting a series of events and a corresponding arrival time for each event;

determining a series of inter-arrivals times relative to a previous event within the series and an associated probability of occurrence of the inter-arrival times relative to a length of a specified time interval;

calculating an expected frequency of events over the series of inter-arrival times for a portion of the fluidic stream having a length equal to the specified time interval, said calculating based on a predetermined distribution;

measuring an observed frequency of events over the series of inter-arrival times within the portion of the fluidic stream; and

generating the sample behavior information based at least in part on a ratio of the observed frequency to the expected frequency of events over the series of inter-arrival times for the portion of the fluidic stream.

5. The method of claim 4, wherein the predetermined distribution includes Poisson distribution.

6. The method of claim 4, wherein the stream is comprised of regularly spaced droplets.

7. The method of claim 4, further comprising:

comparing the entrainment factor to a predetermined value; and

actuating a corrective action in a control system operationally connected to the flow cytometer based on a result of the comparing.

8. The method of claim 7, wherein the corrective action comprises halting the flow of the sample in the flow cytometer.

9. The method of claim 7, further comprising sorting particles in the stream, and wherein the corrective action comprises adjusting the sorting based on the entrainment factor.

11. The method of claim 6, wherein time intervals are measured in fractions of a droplet.

15. A flow system for providing a sample entrainment factor, the system comprising:

a fluidics system comprising a sample tube and a moving fluid column within the sample tube in which particles of a sample move along a common sample path;

a detection system for collecting a signal from each particle as it passes one or more detection stations along the common sample path, each signal being assigned a signal value to form a data point for each particle, the detection system collecting a succession of such data points in a first time interval; and

a control system operationally associated with the fluidics system, the control system configured to;

generate a calculated signal frequency for at least a portion of the first time interval based on a Poisson distribution and the number of data points collected by the detection system during the first time interval; and

generate an experimental signal frequency based on the number of data points in the portion of the first time interval;

compare the experimental signal frequency with that of a calculated signal frequency or a predetermined signal frequency; and

provide an entrainment factor for the sample, said entrainment factor based on said comparing.

16. The system of claim 15, wherein comparing the experimental signal frequency comprising determining a ratio of the experimental signal frequency to that of the calculated signal frequency or the predetermined signal frequency.

17. The system of claim 15, wherein upon determining that the entrainment factor deviates from a predetermined value, the control system actuates a corrective action.

18. The system of claim 17, wherein the corrective action includes one or more of:

interrupting collection of signals by the detection system;

purging the sample tube by said fluidics system;

altering collection of signals via acoustic focusing based on the entrainment factor; and

resuming collection of the signals by the detection system.

19. The system of claim 18, wherein resuming collection of the signals occurs after initiating dispersion of the sample.

20. The system of claim 15, wherein the control system includes a first sorting configuration providing a high collection yield and a second sorting configuration providing a high collection purity, and wherein the control system is configured to select one of the first sorting configuration or the second sorting configuration based on the entrainment factor.