SOME FACTORS AFFECTING THE PRESERVATION OF THE CYTOSKELETON OF CELLS IN CULTURE FOR SCANNING ELECTRON-MICROSCOPY

  • 1 January 1983
    • journal article
    • research article
    • p. 351-359
Abstract
The effects of the composition of the detergent extraction medium on the morphological and molecular preservation of the cytoskeleton of Balb/c 3T3 cells as visualized by scanning electron microscopy and SDS-PAGE [sodium dodecyl sulfate-polyacrylamide gel electrophoresis] are reported. Specific variables tested are the concentrations of Na, K, Ca and Mg ions, and the osmolarity of the medium. The ionic composition of the medium may be more important than osmolarity in determining the composition of the cytoskeleton obtained with Triton X-100 detergent. Ca is particularly deleterious to morphological preservation, and better preservation is obtained when K is the principal cation than when Na is. The best preservation of cytoskeletal structures and the greatest molecular complexity was obtained by extracting in a buffer composed of 1 mM EGTA [ethylene glycol bis(.beta.-aminoethyl ether) N,N''-tetraacetic acid], 4% polyethylene glycol 6000, 1 mM GTP and 100 mM PIPES [piperazine-N,N''-bis(2-ethanesulfonic acid)], pH 6.9.