Actin protein (rhodamine, rabbit skeletal muscle)

Question 1: What is the best way to store actin proteins to insure maximum stability and shelf-life?

Answer 1: Cytoskeleton provides all actin proteins as lyophilized powders so that they can be shipped at room temperature. Upon receipt, the lyophilized powders should be stored at 4°C in a sealed container with desiccant. It is important to monitor the freshness of the desiccant and insure that it continues to absorb moisture to protect the lyophilized actins. With proper storage, the lyophilized actins are guaranteed to be stable for 6 months. Alternatively, actins can be immediately resuspended at the concentration recommended, aliquoted, snap-frozen in liquid nitrogen and stored at -70°C. When thawing frozen aliquots, it is important to thaw rapidly in a room temperature water bath.

Question 2: What is the best way to store F-actin after polymerizing?

Answer 2: G-actin is stable for two days at 4°C and requires a divalent cation, pH 6.5 - 8.0 and ATP for stability. F-actin is stable and can be stored at 4°C for 1-2 weeks. F-actin requires ATP (0.2 mM) and Mg2+ (2 mM) for stability and is unstable below pH 6.5 and above pH 8.5. F-actin is not stable to freezing. F-actin can be transferred to a variety of buffers (e.g. HEPES, phosphate, etc) without detrimental effects. We recommend the addition of antibacterial agents such as 100 &mu, g/ml ampicillin and 10 &mu, g/ml chloramphenicol when storing F-actin at 4°C.

Question 3: Filters for visualizing rhodamine signal?

Answer 3: Theexcitation filter should be set at 535 nm and the emission filter at 585 nm.