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The colchicine-binding protein in procine-brain tissue is a dimer of molecular weight 110,000 that is believed to be the subunit of neuronal microtubules. Conditions are established under which the dimers aggregate with reproducible kinetics. This aggregation reaction, which is monitored by development of turbidity, has the following characteristics: (a) Colchicine inhibits development of turbidity; (b) the reaction inhibited by colchicine is reversed by long-wave ultraviolet irradiation; (c) the aggregation is temperature-dependent; (d) the reaction is nucleotide triphosphate-specific, being stimulated by 1 mM GTP; (e) the reaction appears to be specific for microtubule subunits since in the presence of other added proteins and in curde cell extracts, only microtubule subunits aggregate. On the basis of these criteria, we conclude that we have established an in vitro system for the aggregation of microtubule subunits that shares some of the properties characteristic of the in vivo assembly of cytoplasmic and spindle microtubules.

The addition of vinblastine to high-speed supernatants derived from homogenates of cultured mouse neuroblastoma cells results in the formation of a precipitate which has been characterized as microtubule protein by the following criteria: colchicine-binding activity, molecular weight, amino acid composition, and electrophoretic mobility. The method therefore permits the rapid isolation of microtubule protein from crude supernatants of neuroblastoma cells.