Purification and properties of avian and mammalian filamins.
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This chapter presents properties and procedure for purification of Avian and mammalian filamins. Filamin can be purified from both smooth muscle and nonmuscle cells. However, chicken gizzard is a particularly convenient source because large quantities of fresh tissue are readily obtainable and inexpensive. Two general problems are encountered during filamin purification: proteolytic degradation, and aggregation of the protein. To reduce proteolysis, we routinely work with fresh tissues, carry out all steps at 04 , move as expeditiously as possible through the early steps of purification, and include EDTA in all buffers. Aggregation can be minimized also by working with fresh tissues in the cold, by including reducing agents in the buffers, and by keeping the ionic strength of buffers above 100 mM KCI until all the actin is removed from the preparation. Purification of filamin is monitored by SDS-polyacrylamide gel electrophoresis on 5% linear polyacrylamide slab gels. 1982, Elsevier Inc. All rights reserved.