Phase-correction of radiofrequency-multiplexed signals
Aspects of the present disclosure include methods for characterizing particles of a sample in a flow stream. Methods according to certain embodiments include generating frequency-encoded fluorescence data from a particle of a sample in a flow stream; and calculating phase-corrected spatial data of the particle by performing a transform of the frequency-encoded fluorescence data with a phase correction component. In certain embodiments, methods include generating an image of the particle in the flow stream based on the phase-corrected spatial data. Systems having a processor with memory operably coupled to the processor having instructions stored thereon, which when executed by the processor, cause the processor to calculate phase-corrected spatial data from frequency-encoded fluorescence data of a particle a flow stream are also described. Integrated circuit devices (e.g., field programmable gate arrays) having programming for practicing the subject methods are also provided.
1. A system comprising:
a light source configured to irradiate a sample comprising particles in a flow stream;
a light detection system; and
a processor comprising memory operably coupled to the processor wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to:
generate frequency-encoded fluorescence data from a particle in the flow stream; and
calculate phased-corrected spatial data of the particle by performing a transform of the frequency-encoded fluorescence data with a phase correction component.
2. The system according to claim 1 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to perform a Fourier transform of the frequency-encoded fluorescence data with the phase correction component to generate the phase-corrected spatial data of the particle.
3. The system according to claim 2 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to perform a discrete Fourier transform of the frequency-encoded fluorescence data with the phase correction component to generate the phase-corrected spatial data of the particle.
4. The system according to claim 2 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to perform a short time Fourier transform (STFT) of the frequency-encoded fluorescence data with the phase correction component.
5. The system according to claim 2 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to calculate the phase-corrected spatial data with a digital lock-in amplifier to heterodyne and de-multiplex the frequency-encoded fluorescence data.
6. The system according to claim 1 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to transform the frequency-encoded fluorescence data into the spatial data with a phase correction component that comprises modified transform coefficients.
7. The system according to claim 6 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to calculate a phase correction component that comprises a first phase adjustment and a second phase adjustment.
8. The system according to claim 7 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to calculate the first phase adjustment by:
multiplying an output signal from a brightfield photo photodetector with a predetermined constant signal to produce a phase adjustment value; and
calculating the arctangent of the phase adjustment value to generate the first phase adjustment.
9. The system according to claim 8 , wherein the phase adjustment value is a sum of all bins in a discrete Fourier transform of the frequency-encoded fluorescence data.
10. The system according to claim 7 , wherein the first phase adjustment is an interferometric phase adjustment.
11. The system according to claim 1 , wherein the light detection system comprises a photodetector configured to detect one or more of light absorption, light scatter and fluorescence.
12. The system according to claim 11 , wherein the light detection system comprises a brightfield photodetector.
13. The system according to claim 1 , wherein the light detection system comprises a fluorescence detector.
14. The system according to claim 1 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to calculate the second phase adjustment based on a fluorescence lifetime of a fluorophore in the sample.
15. The system according to claim 1 , wherein the system is a flow cytometer.
16. The system according to claim 1 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to generate an image of the particle from the phase-corrected spatial data.
17. The system according to claim 16 , wherein the memory comprises instructions stored thereon, which when executed by the processor, cause the processor to generate an image mask of the particle.
18. The system according to claim 1 , further comprising a cell sorting component configured to sort cells in the sample based on the calculated phase-corrected spatial data.
19. The system according to claim 18 , wherein the cell sorting component comprises a droplet deflector.
20. A system comprising:
a light source configured to irradiate a sample comprising particles in a flow stream;
a light detection system; and
an integrated circuit component programmed for:
generating the frequency-encoded fluorescence data from the particle of a sample in the flow stream; and
calculating phase-corrected spatial data of the particle by performing a transform of the frequency-encoded fluorescence data with a phase correction component.