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Ratiometric pH measurements using LysoSensor DND-192.

Jung Sook Kang1, Yordan Kostov

  • 1Department of Oral Biochemistry and Molecular Biology, College of Dentistry and Research Institute for Oral Biotechnology, Pusan National University, Pusan 602-739, Korea. jsokang@pusan.ac.kr

Journal of Biochemistry and Molecular Biology
|September 26, 2002
PubMed
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This study introduces a ratiometric pH sensing method using LysoSensor DND-192, effective in the physiological pH range. The technique utilizes two fluorophores with distinct lifetimes, enabling accurate measurements even with overlapping emission spectra and strong ambient light.

Area of Science:

  • Biomedical Engineering
  • Analytical Chemistry
  • Cell Biology

Background:

  • Accurate pH monitoring is crucial for biological and chemical processes.
  • Existing ratiometric pH sensing methods often face challenges with spectral overlap and ambient light interference.
  • LysoSensor DND-192 offers potential for advanced pH sensing applications.

Purpose of the Study:

  • To develop and present a novel ratiometric pH sensing method utilizing LysoSensor DND-192.
  • To demonstrate the method's efficacy within the physiological pH range.
  • To overcome limitations of spectral overlap and ambient light in pH measurements.

Main Methods:

  • Employed a ratiometric approach based on two fluorophores with significantly different fluorescence lifetimes.

Related Experiment Videos

  • Utilized frequency discrimination for separating fluorophore emissions, allowing for spectral overlap.
  • Implemented simple long-pass or combined long-/short-pass filters, avoiding narrow-bandpass devices.
  • Conducted measurements under strong ambient light conditions.

    • Main Results:

    • The developed method provides ratiometric pH sensing within the physiological pH range.
    • Successful discrimination of emissions from two fluorophores was achieved through frequency differences.
    • The use of simple filters proved effective despite significant overlap in emission spectra.
    • Robust measurements were obtained even under strong ambient light.

    Conclusions:

    • The presented ratiometric pH sensing method is reliable and effective for physiological pH ranges.
    • This technique offers a robust solution for pH monitoring in challenging environments, including strong ambient light.
    • The method holds broad applicability in intracellular measurements, microscopy, and bioprocess monitoring.