Automated and accurate drop delay for flow cytometry
Disclosed is an automated method and apparatus for automatically setting a drop delay period by detecting calibration particles in a waste stream. The drop delay is incremented over a series of drop delays and the number of calibration particles in the waste stream is detected for each drop delay. The drop delay is selected which has the least number of calibration particles in the waste stream.
1. A flow cytometry system for selecting a desired drop delay for a flow cytometer, the flow cytometer comprising:
means for forming a stream such that the stream forms droplets at a droplet separation point;
means for interrogating the stream at an interrogation point upstream of the droplet separation point, wherein the desired drop delay is representative of the elapsed time between when particles pass through the interrogation point and when a droplet containing the particles separates from the stream at the droplet separation point;
means for charging the droplets;
means for electrostatically deflecting the droplets;
waste tubing configured to form a waste stream that is generated from droplets that are not deflected by the means for electrostatically deflecting the droplets;
a waste stream optical source configured to illuminate the waste stream in the waste tubing;
a detector configured to sense scattered light or fluorescent emissions from calibration particles in the waste stream; and
a controller that is configured to:
operate the flow cytometer for a plurality of drop delay periods while calibration particles are present in the stream, each drop delay period having a different selected drop delay from a range of potential selected drop delays,
cause the means for charging to charge the droplets during each drop delay period according to the selected drop delay for that drop delay period,
cause the means for electrostatically deflecting the droplets to deflect the charged droplets during each of the drop delay periods,
determine information indicative of a rate at which the calibration particles enter the waste stream during each drop delay period based on the sensed scattered light or fluorescent emissions from the calibration particles in the waste stream, and
determine the desired drop delay based upon the information indicative of the rate at which the calibration particles enter the waste stream during each drop delay period.
2. The system of claim 1 , wherein the controller is further configured to set the selected drop delay of the flow cytometer to the desired drop delay.
3. The system of claim 1 , further comprising a pump configured to draw the waste stream through the waste tubing at a flow rate greater than a flow rate at which the stream is flowed.
4. The system of claim 1 , wherein the waste tubing has a larger internal diameter than an exit orifice diameter of a nozzle of the means for forming the stream.
5. The system of claim 1 , wherein the controller determines the desired drop delay using pattern recognition techniques.
6. The system of claim 1 , wherein the detector is also used to detect the calibration particles at the interrogation point in the stream.
7. The system of claim 6 , further comprising:
one or more optical collectors aligned to collect said the scattered light or fluorescent emissions from the calibration particles in the waste stream; and
one or more fiber optic cables aligned to transmit the scattered light or fluorescent emissions to the detector.
8. The system of claim 1 , further comprising:
a display that visually displays the amount of calibration particles in the waste stream for the each drop delay period to allow an operator of the flow cytometer to select a drop delay period.