Image of ActinRed™ 555 ReadyProbes® Reagent packaging

F-actin stain for fixed cells

ActinRed™ 555 ReadyProbes® Reagent is a selective, high-affinity F-actin probe conjugated to the red-orange–fluorescent dye tetramethylrhodamine (TRITC). Like other ReadyProbes® reagents, ActinRed™ 555 reagent is formulated as a room temperature–stable solution and comes in a convenient dropper bottle

This protocol can be used for:

  • F-actin staining in fluorescence microscopy

This protocol should not be used for:

  • Flow cytometry

You will need the following for this protocol:

Protocol

Labeling fixed cells

First, fix and permeabilize cultured cells with a protocol appropriate for your sample. Counterstain as desired. DAPI may be added directly to fixative.

1. Wash the cells 1–3 times in PBS as needed.
2. Add two drops of ActinGreen™ 488 ReadyProbes® Reagent per milliliter of medium.
3. Incubate for 30 minutes, protected from light.
4. Remove the stain solution.
5. Wash the cells 2–3 times in PBS.
6. Image the cells.
Spectral information and storage
  ActinRed™ 555
Excitation/Emission (nm) 540/565
Standard filter set TRITC
EVOS® Light Cube TRITC
Storage conditions Room temperature

 

 



Protocol tips

Multicolor staining of BPAE cells
Multicolor staining of BPAE cells. BPAE cells were fixed and permeabilized using the Image-iT® Fixation/Permeabilization Kit. Actin was stained using ActinRed™ 555 ReadyProbes® Reagent, and nuclei were stained with NucBlue® Fixed Cell ReadyProbes® Reagent.