Actin protein (human platelet non-muscle >99% pure)

Question 1: Do you have pyrene-labeled non-muscle actin for use in a polymerization assay?

Answer 1: Pyrene-labeled non-muscle actin has been shown to be unstable under normal storage conditions and was discontinued. To examine the polymerization of unlabeled non-muscle actin, please click here for a polymerization protocol that uses an excess of unlabeled non-muscle actin (Cat. #APHL99) + a small amount of pyrene-labeled muscle actin (Cat. #AP05). The pyrene muscle actin will not polymerize efficiently on its own at the concentration used in this assay, so the reaction is dependent on unlabeled actin polymerization for F-actin formation. In this way, the pyrene-labeled muscle actin is taken up and polymerized to serve as a reporter for polymerization of the unlabeled non-muscle actin that is present at a much greater concentration.

Question 2: Are the actin products shipped as pure G-actin or a mixture of G- and F-actin?

Answer 2: Most of our actin proteins are sold in the monomer form (G-actin) because this is stable to freezing and lyophilization. That being said, on the day of the experiment, we do recommend incubating the actin on ice for 60 min before beginning the experiment to depolymerize any actin oligomers that might have formed during storage. Typically actin is first diluted to 0.4 or 0.2 mg/ml concentration and then this can be incubated on ice for 60 min to depolymerize any actin oligomers that might have formed. If you are working with an actin concentration above 0.4 mg/ml, we recommend the ice incubation followed by a high-speed centrifugation (100, 000 x g) for 60 min to pellet any actin oligomers that may not have depoymerized. Remove the top 80% of the supernatant and use this as your G-actin stock. We also provide pre-formed actin filaments (CAt. #AKF99) that are shipped lyophilized and upon resuspension, the filaments are ready for use and average 5-10 microns in length.