Systems and method for correction of positionally dependent electromagnetic radiation detected from objects within a fluid column
A discrimination system that forms a fluid column and interrogates objects within the fluid column with an excitation source. An optical arrangement collects output electromagnetic radiation emanating from the excited objects disposed within the fluid column and directs the output electromagnetic radiation to a detector. An analyzer reduces the positional dependency of the detected intensity by normalizing the value based on the position of each object.
1. A discrimination system, comprising:
collection optics that collect output electromagnetic radiation from X-chromosome bearing sperm and Y-chromosome bearing sperm within a measurement region;
a detector that generates an electrical signal responsive to the intensity of the output electromagnetic radiation collected by the collection optics; and
an analyzer having instructions stored thereon i) to normalize the intensity of the output electromagnetic radiation represented in the electrical signal based on the position of the sperm cells in the measurement region, and ii) to discriminate between X-chromosome bearing sperm and Y-chromosome bearing sperm.
2. The discriminating system of claim 1 , wherein the detector comprises a first detector and wherein the system further comprises a second detector that detects the position of the sperm cells in the measurement region.
3. The discriminating system of claim 2 , wherein the second detector comprises a position detector.
4. The discriminating system of claim 3 , wherein the position detector is one selected from the group of: a camera, a CCD, PSD, SiPM split detector, and photodiode array.
5. The discrimination system of claim 1 , wherein the detector comprises a first detector that: i) generates an electrical signal responsive to the intensity of the output electromagnetic radiation collected by the collection optics and ii) detects the position of sperm cells in the measurement region.
6. The discrimination system of claim 5 , wherein the first detector is not located in an image plane of the collection optics.
7. The discrimination system of claim 5 , wherein the first detector comprises one selected from the group consisting of a: PMT split detector, SiPM split detector, and photodiode array, an array of PMTs, and an array of SiPMs.
8. The discrimination system of claim 1 , wherein the instructions stored on the analyzer for normalizing the intensity of the output electromagnetic radiation represented in the electrical signal based on the position of the sperm cells in the measurement region applies a correction to the intensity of the output electromagnetic radiation represented in the electrical signal.
9. The discrimination system of claim 8 , wherein the amount of correction is determined for each position on a major axis of a core stream at the measurement region, and wherein the correction is applied to the electrical signal representing each sperm cell based on its detected position on the major axis of the core stream.
10. The discriminating system of claim 1 , wherein the analyzer includes instructions for discriminating viable X-chromosome bearing sperm or viable Y-chromosome bearing sperm from other cells.
11. The discriminating system of claim 1 , wherein the sperm cells located within an elliptical column of sample fluid, where the sample fluid is formed coaxially with an outer layer of sheath fluid and the sheath fluid comprises a generally cylindrical shape.
12. The discriminating system of claim 1 , further comprising an element that modifies the output electromagnetic radiation to increase the uniformity of output electromagnetic radiation collected by the collection optics of sperm cells at different positions.
13. A method, comprising:
collecting output electromagnetic radiation from X-chromosome bearing sperm and Y-chromosome bearing sperm within a measurement region;
generating an electrical signal responsive to the intensity of the collected output electromagnetic radiation;
normalizing the intensity of the output electromagnetic radiation represented in the electrical signal based on the position of the sperm cells in the measurement region, and
discriminating between X-chromosome bearing sperm and Y-chromosome bearing sperm.
14. The method of claim 13 , further comprising detecting the position of sperm cells in the measurement region.
15. The method of claim 14 , wherein the analyzer includes instructions for discriminating for viable X-chromosome bearing sperm or viable Y-chromosome bearing sperm.
16. The method of claim 13 , wherein the sperm cells in measurement region are located in a generally elliptical cross section of core stream contained within a generally circular cross section of sheath fluid, and wherein sperm cells in the fluid column are located at differing positions on a major axis of the generally elliptical cross section of the core stream.
17. The method of claim 16 further comprising applying a correction to the intensity of the output electromagnetic radiation represented in the electrical signal.
18. The method of claim 17 , further comprising determining a correction factor for each position on the major axis of the core stream and applying the correction factor to each event based on the position on the major axis of the core stream.
19. The method of claim 13 , further comprising an element that modifies the output electromagnetic radiation to increase the uniformity of output electromagnetic radiation collected from sperm cells at different positions.