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POLYMERIZING ACTIN

Materials:
  • Ultra-Pure Actin (Cat. No. 160101) or Pure Actin (Cat. No. 190101)
  • Actin Working Buffer (Cat. No. 000102; 5 mM Tris-HCl and 0.2 mM CaCl2, pH 8.0)
  • Actin Polymer Buffer (10X; Cat. No. 000103; 500 mM KCl and 20 mM MgCl2)
  • ATP
  • DTT
Equipment:
  •  Microcentrifuge at 4°C
  •  Spectrophotometer
Technical Notes:
  • Avoid diluting actin beyond its critical concentration (dependent on ionic strength)
  • Polymerized actin (F-actin) is stable at 4°C for several weeks, do not freeze
Protocol:
1. Reconstitute actin to 0.4 mg/ml (see respective protocol for details)

 

2. Assemble the Polymerization Reaction

a. Add 1/10 volume Actin Polymer Buffer to actin

b. Add ATP to 1 mM


3. Polymerize

a. Incubate at room temperature for 1 hour