Cell Staining: Fluorescent Labelling of Endocytic Compartments


Endocytic compartments are acidic organelles involved in the transport of extracellular solutes and ligands from the plasma membrane to early, then to late endosomes and eventually to lysosomes. Endocytic compartments can be fluorescently labelled by fluid phase endocytosis, internalized specific ligands, resident proteins and pH‐sensitive probes.

Keywords: endocytosis; endosome; lysosome; membrane traffic; green fluorescent protein

Figure 1.

Schematic view of the endocytic pathway. Ligands and solutes are internalized from the external medium and reach the early endosomes after 5–10 min. Materials can be recycled to the plasma membrane through the recycling endosomes or subsequently transferred to late endosomes (15–30 min) and eventually to lysosomes (40–60 min) for degradation. Stationary endocytic organelles can be identified by specific markers: (1) EEA1, Rab5, the transferrin receptor for early endosomes; (2) Rab4 and the transferrin receptor for the recycling compartment; (3) Rab7, the cation‐independent mannose 6‐phosphate receptor (CI‐MPR) for late endosomes; (4) lysosomal hydrolases such as cathepsin D and lysosomal membrane glycoproteins (LAMP1, LAMP2, CD63), vacuolar ATPase for lysosomes. A lipid, the lysobisphosphatidic acid (LBPA), has been described to be enriched in late endocytic compartments (Kobayashi et al., ).

Figure 2.

Examples of fluorescent labelling of endocytic compartments. (a) Fluorescent labelling of early and late endosomes. HeLa cells were allowed to internalize fluorescein isothiocyanate (FITC)‐transferrin (green), fixed/permeabilized and then incubated with an anti‐Rab7 antibody revealed by a Texas red‐conjugated secondary antibody (red). The dotted line delimits the nucleus (N). (b) LAMP1‐GFP is recruited on vacuoles surrounding intracellular bacteria (arrowheads). Live HeLa cells expressing LAMP1‐GFP were infected with Salmonella typhimurium and directly observed under an epifluorescence microscope. (c) In vitro reconstitution of early endosome fusion. Isolated early endosomes containing FITC‐BSA (green) or rhodamine‐BSA (red) were mixed in a fusion‐competent assay. The yellow colour results from the mixing of contents of both fluorescent probes. The scale bars represent (a) 10 μm, (b) 5 μm and (c) 1 μm.



Brandtzaeg P (1973) Conjugates of immunoglobulin G with different fluorochromes. Scandinavian Journal of Immunology 2(3): 273–290.

Bucci C, Thomsen P, Nicoziani P, McCarthy J and van Deurs B (2000) Rab7: a key to lysosome biogenesis. Molecular Biology of the Cell 11(2): 467–480.

Chalfie M, Tu Y, Euskirchen G, Ward WW and Prasher DC (1994) Green fluorescent protein as a marker for gene expression. Science 263(5148): 802–805.

Chavrier P, Van der Sluijs P, Mishal Z, Nagelkerken B and Gorvel JP (1997) Early endosome membrane dynamics characterized by flow cytometry. Cytometry 29(1): 41–49.

Dautry‐Varsat A, Ciechanover A and Lodish HF (1983) pH and the recycling transferrin during receptor‐mediated endocytosis. Proceedings of the National Academy of Sciences of the USA 80(8): 2258–2262.

Kobayashi T, Stang E, Fang KS et al. (1998) A lipid associated with the antiphospholipid syndrome regulates endosome structure and function. Nature 392(6672): 193–197.

Matz MV, Fradkov AF, Labas YA et al. (1999) Fluorescent proteins from nonbioluminescent Anthozoa species. Nature Biotechnology 17(10): 969–973.

Pizarro‐Cerda J, Méresse S, Parton R et al. (1998) Brucella abortus intracellular pathway in HeLa cells: transit through the autophagic pathway and replication in the endoplasmic reticulum. Infection and Immunity 66(12): 5711–5724.

Tooze J and Hollinshead M (1991) Tubular early endosomal networks in AtT20 and other cells. Journal of Cell Biology 115(3): 635–653.

Wubbolts R, Fernandez‐Borja M, Oomen L et al. (1996) Direct vesicular transcript of MHC Class II molecules from lysosomal structures to the cell surface. Journal of Cell Biology 135(3): 611–622.

Further Reading

Gruenberg J and Maxfield FR (1995) Membrane transport in the endocytic pathway. Current Opinion in Cell Biology 7(4): 552–563.

Ellenberg J, Lippincott‐Schwartz J and Presley JF (1999) Dual‐colour imaging with GFP variants. Trends in Cell Biology 9(2): 52–56.

Mellmann I (1996) Endocytosis and molecular sorting. Annual Review of Cell and Developmental Biology 12: 575–625.

Wilson T and Hastings JW (1998) Bioluminescence. Annual Review of Cell and Developmental Biology 14: 197–230.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Gorvel, Jean‐Pierre, and Méresse, Stéphane(Apr 2001) Cell Staining: Fluorescent Labelling of Endocytic Compartments. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1038/npg.els.0002630]