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 method comprising:
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.
2. The method according to claim 1 , wherein the phase-corrected spatial data is calculated by performing a Fourier transform of the frequency-encoded fluorescence data with the phase correction component.
3. The method according to claim 2 , wherein the phase-corrected spatial data is calculated by performing a discrete Fourier transform of the frequency-encoded fluorescence data with the phase correction component.
4. The method according to claim 2 , wherein the phase-corrected spatial data is calculated by performing a short time Fourier transform (STFT) of the frequency-encoded fluorescence data with the phase correction component.
5. The method according to claim 1 , wherein the phase-corrected spatial data is calculated with a digital lock-in amplifier to heterodyne and de-multiplex the frequency-encoded fluorescence data.
6. The method according to claim 1 , wherein the phase correction component comprises modified transform coefficients that are used to transform the frequency-encoded fluorescence data into the phase-corrected spatial data.
7. The method according to claim 1 , wherein generating the frequency-encoded fluorescence data from the particle comprises detecting light from the particle in the sample with a light detection system.
8. The method according to claim 7 , wherein the light detected from the particle comprises light absorption, light scatter, emitted light or a combination thereof.
9. The method according to claim 1 , wherein the phase correction component comprises a first phase adjustment and a second phase adjustment.
10. The method according to claim 9 , wherein the first phase adjustment comprises an output signal from the light detection system.
11. The method according to claim 10 , wherein the first phase adjustment comprises an output signal from a brightfield photodetector.
12. The method according to claim 11 , further comprising calculating the first phase adjustment by:
multiplying an output signal from the brightfield 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.
13. The method according to claim 12 , wherein the phase adjustment value is a sum of all bins in a discrete Fourier transform of the frequency-encoded fluorescence data.
14. The method according to claim 9 , wherein the first phase adjustment is an interferometric phase adjustment.
15. The method according to claim 9 , further comprising calculating the second phase adjustment based on a fluorescence lifetime of a fluorophore in the sample.
16. The method according to claim 1 , wherein the phase-corrected spatial data of the particle is calculated from the frequency-encoded fluorescence data by an integrated circuit device.
17. The method according to claim 1 , further comprising generating an image of the particle from the phase-corrected spatial data.
18. The method according to claim 1 , further comprising sorting the particle.
19. The method according to claim 1 , further comprising irradiating the flow stream with a light source.
20. The method according to claim 19 , wherein the method comprises irradiating the flow stream with a first beam of frequency shifted light and second beam of frequency shifted light.
21. The method according to clause 20 , wherein the first beam of frequency shifted light comprises a local oscillator (LO) beam and the second beam of frequency shifted light comprises a radiofrequency comb beam.
22. The method according to claim 20 , further comprising:
applying a radiofrequency drive signal to an acousto-optic device; and
irradiating the acousto-optic device with a laser to generate the first beam of frequency shifted light and the second beam of frequency shifted light.