• Study the relationship between signal transduction and actin dynamics
• Study in vivo actin-binding protein (ABP) effects on actin dynamics
• Follow G:F actin ratio dynamics throughout morphological changes, such as development
• Study the effects of pharmaceutical compounds on the ratio of G:F actin

Kit contents (sufficient for 100 x 1 ml lysates)

• Lysis and F-actin stabilization buffer
• Protease inhibitor cocktail
• ATP
• F-actin depolymerizing buffer
• F-actin enhancing solution
• G-actin protein standard
• Anti-actin mAb
• SDS sample buffer
• DMSO

Equipment & materials required

• Temperature-controlled ultra centrifuge capable of reaching 100,000 x g. Accepts 100 µl sample volumes. The assay can be adapted for larger volumes; however, this will result in fewer assays per kit.
• Small homogenizer (low ml volumes) or 25G needle & syringe
• SDS-PAGE and Western blot equipment
• Anti-mouse secondary antibody

This kit enables in vivo measurement of the Filamentous: Globular (F: G) actin ratio in cells and tissues, providing insight into actin dynamics under set experimental conditions. Cells are lysed in a detergent-based buffer that preserves both G- and F-actin. The buffer solubilizes G-actin, while F-actin remains insoluble. Centrifugation separates the fractions, pelleting F-actin and leaving G-actin in the supernatant. Both fractions are analyzed by SDS-PAGE and quantified via Western blot.

Key characteristics

• Semi-quantitative: accurately measures the in vivo F: G actin ratio
• Separation process: simple ultracentrifugation step
• Detection method: SDS-PAGE or Western blot
• Timeframe: Approximately 8h, typically performed over two days