Product Uses Include
- To show quantitative / qualitative effects on actin polymerization by the addition of a tissue extract, an actin binding protein or compound.
- To show quantitative / qualitative effects on actin polymerization by addition of an F-actin nucleating protein, compound or extract.
- To show quantitative / qualitative effects on steady-state F-actin levels by addition of an F-actin severing protein, compound or tissue extract.
- To show quantitative / qualitative effects on actin depolymerization by addition of an actin binding protein, compound or tissue extract.
Introduction
The Actin Polymerization Biochem Kit™ is based on the enhanced fluorescence of pyrene conjugated actin that occurs during polymerization. The enhanced fluorescence that occurs when pyrene G-actin (monomer) forms pyrene F-actin can be measured in a fluorimeter to follow polymerization over time. Also, by using preformed pyrene F-actin, it is possible to follow depolymerization. Both cell/tissue extracts and purified proteins can be added to the reaction mixture to identify their effect on actin polymerization. The components of the kit can also be used separately for other actin based assays such as a spin-down assays to detect F-actin binding proteins (see also BK001) or size exclusion chromatography to identify G-actin binding proteins.
While this kit comes with pyrene labeled skeletal muscle actin, it can also be used to study polymerization of other types of actin such as non-muscle actin (Cat. # APHL99) or cardiac actin (Cat. # AD99). Polymerization assays with these actins can be performed using a 10:1 ratio between the actin you want to study and the included pyrene actin
Kit contents
The kit contains enough materials for 30-100 assays depending on assay volume. The following reagents are included:
- 5 x 1 mg Pyrene labeled actin (Cat. # AP05).
- General Actin Buffer (Cat. # BSA01).
- Actin Polymerization Buffer (Cat. # BSA02).
- ATP 100mM (Cat. # BSA04).
- Tris-HCl pH 7.5, 100 mM
- Manual with detailed protocols and extensive troubleshooting guide.
Equipment needed
- Fluorescence spectrophotometer (cuvette or 96-well plate) with 4-10 nm bandwidth at 365 nm excitation wavelength, and 4-10 nm bandwidth at 407 nm emission wavelength.
- Small capacity (100-1000 µl) fluorescence spectrophotometer cuvette or 96-well plate.
