As is well known, the actin cytoskeleton is a highly dynamic structure, involved in a large number of cellular processes, including muscle contraction, lamellopodial extrusion, cell locomotion, cytokinesis, intracellular transport and many more. Under the tight regulation of more than 50 actin binding proteins (ABPs) and in response to a wide variety of internal and external stimuli, the morphology of the actin cytoskeleton changes rapidly.

For more than 20 years now, Cytoskeleton Inc. has provided a large selection of robust and user friendly solutions (pure and active actin proteins/Biochemical assays and actin staining probes..) which have helped many researchers to solve fundamental questions in cell signalling, cytoskeleton and actin research fields.

For more than 2 years now, the Silicon Rhodamine-like (SiR) technology has allowed the live cell imaging field with fluorescence microscopy to evolve significantly.

Fluorescent SiR-probes have appeared as the best alternative tools for studying Actin (SiR-actin), Microtubules (SiR-Tubulin), DNA (SiR-DNA) and now lysosome (SiR-Lysosome) for live cell imaging. Who better to show this? Well, here’s how other researchers have been using them to get optimal results.

Today, I’d like to give you an overview about methods in actin research with validated R&D products and kits which will allow you to measure binding to actin and effects on the polymerisation dynamics of actin.

Actin functions as one of the major cytoskeleton structures. It is involved in a plethora of processes in cell biology:  stabilizing the cell shape, cell movements (e.g. cell migration)  and intracellular movements and transport mechanisms.

Actin is a 43 kDa protein that is very highly conserved between species. Actin has three main isotypes (α-actin, β-actin and γ-actin), which show >90% amino-acid (aa) homology between isotypes and >98% homology within members of a particular isotypic group.

The Silicon Rhodamine-like (SiR) technology has significantly contributed to the recent development of DNA and cytoskeletal analysis by live cell imaging.

In 2014, two new Silicon Rhodamine-like (SiR) fluorescent probes were released for studying actin & tubulin by live cell imaging. SiR-Actin and SiR-Tubulin are fluorescent probes compatible with most microscopes (including super-resolution settings) that directly stain actin & tubulin without the need to transfect cells with vectors expressing fluorescently labeled Actin or Tubulin. The two original dyes were successfully followed by a new SiR-DNA probe in order to visualize DNA in living cells.

The existing SiR stains have a λ_abs of 652 nm and a λ_em of 674 nm to be used with the Cy5 filter (Fig 1).

However, the continuously growing number of researchers using these stains asked us whether stains with different biophysical properties would be made available. In other words, they were asking “is there another colour to allow for double staining e.g. of Actin and Tubulin in living cells?“