- Research Article
Monitoring of membrane phospholipid scrambling in human erythrocytes and K562 cells with FM1-43 — a comparison with annexin V-FITC
Cellular & Molecular Biology Letters volume 19, pages 262–276 (2014)
The styryl dye FM1-43 becomes highly fluorescent upon binding to cell membranes. The breakdown of membrane phospholipid asymmetry in ionophore-stimulated T-lymphocytes further increases this fluorescence [Zweifach, 2000]. In this study, the capacity of FM1-43 to monitor membrane phospholipid scrambling was explored using flow cytometry in human erythrocytes and human erythrocyte progenitor K562 cells. The Ca2+-dependent phosphatidylserine-specific probe annexin V-FITC was used for comparison. The presented data show that the loss of phospholipid asymmetry that could be induced in human erythrocytes by elevated intracellular Ca2+ or by structurally different membrane intercalated amphiphilic compounds increases the FM1-43 fluorescence two- to fivefold. The profile of FM1-43 fluorescence for various treatments resembles that of phosphatidylserine exposure reported by annexin V-FITC. FM1-43 detected the onset of scrambling more efficiently than annexin V-FITC. The amphiphile-induced scrambling was shown to be a Ca2+-independent process. Monitoring of scrambling in K562 cells caused by NEM-induced Ca2+-release from intracellular stores and by Ca2+ and ionophore A23187 treatment showed that the increase in FM1-43 fluorescence correlated well with the number of annexin V-FITC-detected phosphatidylserine-positive cells. The results presented here show the usefulness of FM1-43 as a Ca2+-independent marker of dissipation in asymmetric membrane phospholipid distribution induced by various stimuli in both nucleated and non-nucleated cells.
bovine serum albumin
octaethylene glycol mono n-decyl ether
octaethylene glycol mono n-dodecyl ether
4-(6-acetoxymethoxy-2, 7-dichloro-3-oxo-9-xanthenyl)-4′-methyl-2,2′(ethylenedioxy)dianiline-N,N,N′,N′-tetraacetic acid tetrakis (acetoxymethyl) ester
(n-(3-triethylammoniumpropyl)-4-(4-(dibutylamino) styryl) pyridinium dibromide)
phorbol 12-myristate 13-acetate
sodium dodecyl sulphate
sodium tetradecyl sulphate
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Wróbel, A., Bobrowska-Hägerstrand, M., Lindqvist, C. et al. Monitoring of membrane phospholipid scrambling in human erythrocytes and K562 cells with FM1-43 — a comparison with annexin V-FITC. Cell Mol Biol Lett 19, 262–276 (2014). https://doi.org/10.2478/s11658-014-0195-3
- Phospholipid scrambling
- Annexin V-FITC
- Phosphatidylserine exposure
- Human erythrocytes
- K562 cells