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FM4-64, a membrane-selective red fluorescent dye, belongs to the FM family of styrylpyridinium dyes. FM4-64 is widely used to study endocytosis and exocytosis, vesicle trafficking and organelle organization in living animal.

FM4-64 readily translocates to the nuclear envelope (NE) at physiological temperature where it exhibits enhanced fluorescence when excited at 620-650 nm in contrast to 480-520 nm excitation in the endocytic pathway and in the endoplasmic reticulum (ER). In vitro data indicated that the dye reveals an enrichment of negatively charged lipids, presumably due to local phospholipid synthesis. Dual-excitation imaging of FM4-64 in relation to lamina-associated polypeptide-1-green fluorescent protein during mitosis suggested that the disassembly of NE preserves microscale lipid complexes in the ER.
For studies onplant and fungal cells, FM4-64 is usually preferred to FM1-43 due to its superior brightness, greater contrast, higher photostability, and its red-shifted emission facilitating concomitantuse with the green fluorescent protein (GFP).

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