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A Rapid Method for Multispectral Fluorescence Imaging of Frozen Tissue Sections
Published on: March 30, 2020
Multispectral fluorescence imaging to assess pH in biological specimens
Matthew R Hight1, Donald D Nolting, Eliot T McKinley
1Vanderbilt University, Department of Physics, Nashville, Tennessee 37221, USA.
Journal of Biomedical Optics
|February 2, 2011
Summary
This study introduces multispectral fluorescence imaging (MSFI) to measure extracellular pH (pHe) in tumor tissues. The method accurately quantifies pH in ex vivo samples, offering potential for disease research.
Area of Science:
- Biomedical Imaging
- Biophysical Chemistry
- Cancer Research
Background:
- Accurate measurement of tissue pH is crucial for understanding biological processes and diseases like cancer.
- Existing methods for quantifying extracellular pH (pHe) in tissue are limited.
- Multispectral fluorescence imaging (MSFI) offers a potential avenue for quantitative pH assessment.
Purpose of the Study:
- To evaluate multispectral fluorescence imaging (MSFI) for the quantitative measurement of extracellular pH (pHe) in dye-perfused, ex vivo tumor specimens.
- To develop and validate a method for accurate pHe determination using SNARF-4F dye and commercially available imaging instrumentation.
- To explore the feasibility of high-resolution pHe mapping in preclinical cancer models.
Main Methods:
- Utilized SNARF-4F, a pH-sensitive fluorescent dye, with standard fluorimetry to characterize emission properties.
- Determined a correction factor (CF) to account for suboptimal excitation within the imaging system's constraints.
- Applied spectral unmixing of MSFI data from dye solutions and tissue phantoms to quantify protonated and deprotonated dye species.
- Assessed MSFI feasibility for pHe mapping in human colorectal cancer xenografts.
Main Results:
- A correction factor derived from fluorimetry enabled accurate pH determination using MSFI in aqueous solutions and tissue phantoms.
- MSFI successfully quantified equilibrium concentrations of protonated and deprotonated SNARF-4F dye.
- The study demonstrated the suitability of MSFI for quantitative pHe measurement in ex vivo dye-perfused tumor tissue.
- Feasibility for high-resolution pHe mapping in colorectal cancer xenografts was explored.
Conclusions:
- Multispectral fluorescence imaging (MSFI) provides a viable method for quantitative extracellular pH (pHe) measurement in ex vivo, dye-perfused tissues.
- This technique has the potential to advance the study of pH in various preclinical disease models, including cancer.
- The developed correction factor ensures accurate pH quantification despite instrumental limitations.
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