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Calibration beads containing luminescent lanthanide ion complexes
Citation:
Leif, R. C., D. Jin, J. Piper, L. M. Vallarino, J. W. Williams, S. Yang, AND R. M. ZUCKER. Calibration beads containing luminescent lanthanide ion complexes. In Proceedings, SPIE--BIOS, San Jose, CA, January 20 - 26, 2008. SPIE/International Society for Optical Engineering, Bellingham, WA, 17-26, (2008).
Impact/Purpose:
The reliability of lanthanide luminescence measurements, by both flow cytometry and digital microscopy, will be enhanced by the availability of lanthanide calibration beads. These beads can also be used to characterize spectrographic instruments, including microscopes and flow cytometers. The 5 µm beads can be used for spectral calibration of microscopes equipped with a spectrograph and as test particles for time-delayed luminescence flow cytometers. The europium-complex labeled beads provide the great value of having well-defined, reproducible emission peaks and small full widths at half maximum. The narrow band-width and highly reproducible red emissions of these beads are critical for the detection and quantitation of probes (i.e. CY 7 or Draq 5) that fluoresce in the 650–800 nm range
Description:
The reliability of lanthanide luminescence measurements, by both flow cytometry and digital microscopy, will be enhanced by the availability of narrow-band emitting lanthanide calibration beads. These beads can also be used to characterize spectrographic instruments, including microscopes. Methods: 0.5, 3, and 5 micron (m) beads containing a luminescent europium-complex were manufactured and the luminescence distribution of the 5 m beads was measured with a time-delayed luminescence flow cytometer and a timedelayed digital microscope. The distribution of the luminescence intensity from the europium-complex in individual beads was determined on optical sections by confocal microscopy. The emission spectra of the beads under UV excitation were determined with a PARISS spectrophotometer. The kinetics of the luminescence bleaching caused by UV irradiation were measured under LED excitation with a fluorescence microscope. Results: The kinetics of UV bleaching were very similar for the 0.5, 3, and 5 m beads. Emission peaks were found at 592, 616, and 685 nanometers (nm). The width of the principal peak at half-maximum (616 nm) was 9.9 nm. The luminescence lifetimes in water and in air were 340 and 460 microseconds (s), respectively. The distribution of the europium- complex in the beads was homogeneous. Conclusions: The 5 m beads can be used for spectral calibration of microscopes equipped with a spectrograph, as test particles for time-delayed luminescence flow cytometers, and possibly as labels for macromolecules and cells.
