The Pennsylvania Green Fluorophore: a hybrid of Oregon Green and Tokyo Green for the construction of hydrophobic and pH-insensitive molecular probes

[structure: see text] Fluorescent small molecules are powerful tools for exploring cellular biology. As a more hydrophobic, photostable, and less pH-sensitive alternative to fluorescein, we synthesized Pennsylvania Green, a bright, monoanionic fluorophore related to Oregon Green and Tokyo Green. Comparison of membrane probes comprising N-alkyl-3beta-cholesterylamine linked to 4-carboxy-Tokyo Green (pK(a) approximately 6.2) and 4-carboxy-Pennsylvania Green (pK(a) approximately 4.8) revealed that only Pennsylvania Green was highly fluorescent in acidic early and recycling endosomes within living mammalian cells.