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Flow Cytometry Purification of Mouse Meiotic Cells
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Deep ultraviolet lasers for flow cytometry.

William Telford1, Thierry Georges2, Clint Miller3

  • 1Experimental Transplantation and Immunology Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
|November 14, 2018
PubMed
Summary

A new 280 nm deep ultraviolet laser effectively excites quantum dots for flow cytometry. This deep UV laser shows promise for enabling more than 30 simultaneous parameters in high-dimensional analysis.

Keywords:
Qdotbrilliant ultraviolet dyedeep ultraviolet laserflow cytometerquantum nanocrystalsultraviolet laser

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Area of Science:

  • Flow Cytometry
  • Optical Engineering
  • Biotechnology

Background:

  • High-dimensional flow cytometry relies on multiple laser wavelengths for fluorescent probe excitation.
  • Current Brilliant Ultraviolet (BUV) dyes enable up to 30-color analysis, but demand exists for >30 parameters.
  • New deep ultraviolet (UV) polymer dyes are under development for expanded analysis capabilities.

Purpose of the Study:

  • To evaluate a 280 nm deep UV laser as a potential excitation source for novel polymer fluorochromes.
  • To assess the utility of deep UV laser excitation in flow cytometry.
  • To determine spectral compatibility and potential limitations for future high-dimensional cytometry.

Main Methods:

  • A 280 nm deep UV laser was used for excitation.
  • Quantum nanoparticles (Qdots) served as surrogate probes due to unavailability of deep UV polymer dyes.
  • Excitation efficiency and autofluorescence were measured and compared to traditional UV sources.

Main Results:

  • The 280 nm deep UV laser effectively excited Qdots, comparable to traditional UV sources.
  • Deep UV 280 nm excitation showed minimal overlap with BUV dyes, indicating spectral compatibility.
  • Significant autofluorescence was observed in the violet to blue spectrum upon deep UV excitation.

Conclusions:

  • A 280 nm deep UV laser is a viable option for exciting novel polymer dyes in flow cytometry.
  • This laser wavelength supports spectral multiplexing with existing BUV dyes.
  • Future deep UV fluorochrome development must address and mitigate observed autofluorescence.