This work compares the efficacy of varying the concentrations of cryoprotectants when freezing samples of rat muscles for later use in tissue culture. The best yields were obtained with DMSO associated with glycerol and sucrose; before plating, the best results indicated 45% of cells with respect to controls, and delayed (24-36 h) myotube maturation. Maturation was studied by analysing the molecular forms of acetylcholinesterase and the ability of myotubes to form synapses in the presence of neurons, in fresh and frozen muscle.