Invitrogen™
Alexa Fluor™ 546 Phalloidin
A22283
N/A
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To explore the possible reorganization of the actin cytoskeleton as a consequence of cholesterol depletion, it is crucial to quantitatively measure the amount of F-actin under these conditions. I developed a quantitative high-resolution confocal microscopy-based approach that allows measurements of F-actin content using an image reconstruction method. To quantitatively estimate the extent of F-actin reorganization, we treated cells with increasing concentrations of lovastatin (to deplete membrane cholesterol) and labeled F-actin with Alexa Fluor 546 conjugated phalloidin (a specific fluorescent probe for F-actin).
Immunofluorescence
CHO-K1 Cells, Fixed
Final concentration: 0.3 μM in PBS containing 1 mM CaCl 2 and 0.5 mM MgCl 2 for 1 h at room temperature (∼23◦C) in dark
None
Confocal microscopy
Representative confocal micrographs showing the organization of F-actin in control and cells treated with increasing concentrations of lovastatin. F-actin was labeled with Alexa Fluor 546 conjugated phalloidin. The MIPs of 15 z-sections from the base of the coverslip (~4.8 µm from the base into the cell) are shown in the left panels. An increase in F-actin filaments can be observed upon treatment with increasing concentrations of lovastatin. Panels on the right represent the iso-surfaces (defined as voxel contours of equal fluorescence intensity) generated from the z-sections corresponding to MIPs shown in the respective left panels. The scale bars represent 10 µm.
https://doi.org/10.1016/j.jlr.2022.100206
Quick staining and the results are reproducible.
Expensive.
Works great for labeling F-actin in fixed cells.