Quantitation of cap Z in conventional actin preparations and methods for further purification of actin

Gel‐filtration is commonly used to remove contaminants from conventional actin prepared by the method of Spudich and Watt. It has been shown that this procedure removes the majority of a factor that reduces the low‐shear viscosity of actin. We have previously reported that this factor is Cap Z, a barbed end capping protein. We now establish that, even after gel‐filtration, enough Cap Z can be present in conventionally prepared actin to affect events occurring at the barbed ends of actin filaments. We also demonstrate that the concentration of Cap Z can be reduced to more than a log below the K_D for binding of Cap Z to actin by either (1) immunoabsorbtion of conventionally prepared actin with anti‐Cap Z antibodies, or (2) an additional cycle of polymerization/depolymerization followed by repeat gel‐filtration. © 1995 Wiley‐Liss, Inc.