[4][5] However, its adverse characteristics e.g. its cytotoxicity and blue-light instability or low solubility in water often make its application challenging.

[16] Blebbistatin has also been found to stabilize the super relaxed state in the myofilaments, where myosin heads are in a helical order and interact with each other but not with actin.

[7] Reduction of the concentration of blebbistatin to 6.25 uM allows for FRET imaging in isolated adult mouse cardiac muscle cells.

[5] Long-term incubation with blebbistatin results in cell damage and cytotoxicity, which are independent of the myosin inhibitory effect.

The main aims of the structure-activity relationship work on the blebbistatin scaffold are the improvement of the physicochemical properties and the ATPase inhibitory potency, for use as chemical or pharmacological tools.

It has been successfully used in fluorescent imaging experiments involving myosin IIA-GFP expressing live dendritic cells[38] A water-soluble blebbistatin derivative developed in 2016,[9] its high water solubility (~400 uM) enables in vivo research applications.

[39] A permanent inhibition of myosin may be achieved by covalently crosslinking the inhibitor azidoblebbistatin to its target by photoaffinity labeling (PAL).

[44] In research, it is useful compound for control treatment, to check the non-myosin related toxic effects of blebbistatin.

The blebbistatin scaffold has been modified in several ways to optimize myosin isoform specificity or to improve the inhibitory properties and to map the structure-activity relationship.