Intracellular dye heterogeneity determined by fluorescence lifetimes

Area of Science:

• Cell Biology
• Biophysics
• Fluorescence Spectroscopy

Background:

• Cellular localization of fluorescent probes is influenced by dye chemistry and cellular microenvironments.
• Understanding these environments is crucial for accurate biological imaging and analysis.

Purpose of the Study:

• To investigate the number and types of cellular environments within an epithelial cell line.
• To characterize the localization of distinct fluorescent dyes using fluorescence lifetime measurements.

Main Methods:

• Measured fluorescence lifetimes of rhodamine-B, 3,3'-dihexadecylindocarbocyanine-(C3) (diI), and a novel rhodamine-cardiolipin conjugate (Collarein) in Madin-Darby canine kidney (MDCK) cells.
• Analyzed fluorescence decay curves using exponential fitting.
• Correlated lifetime data with fluorescence microscopy observations.

Main Results:

• Rhodamine-B and diI exhibited multi-exponential fluorescence decay, indicating heterogeneous cellular environments.
• Collarein, designed for plasma membrane localization, showed single-exponential decay.
• Microscopy confirmed heterogeneous distribution for diI and rhodamine-B, and surface localization for Collarein.

Conclusions:

• Fluorescence lifetime analysis effectively differentiates cellular environments based on dye localization.
• Collarein serves as a specific probe for the plasma membrane in MDCK cells.
• The study demonstrates the utility of distinct fluorescent probes for mapping cellular compartments.