The effects of colchicine and cytochalasin B (CB) on the ultrastructure, calcium distribution and transport characteristics of the intestine were investigated. Colchicine at low doses inhibited both cellular uptake and transepithelial transport of calcium. At high doses uptake and transport were enhanced. Neither of these effects were dependent on the disruption of microtubules. CB reduced calcium uptake and translocation to the blood but only at relatively high concentrations (greater than 25 ..mu..g/ml). CB treatment (50 ..mu..g/ml) did not disrupt the brush border microfilaments, but rather induced the formation of large vacuoles within the tissue. The site of action of colchicine and CB appears to be at the brush border membrane, and does not appear to involve either microtubules or microfilaments. Both colchicine and CB had a limited effect on the cellular distribution of calcium. A coordinated electron probe-electron microscope study of the intestinal epithelium indicated that relatively high concentrations of calcium were associated with the rough endoplasmic reticulum (and possibly mitochondria), and areas of mucin accumulation in goblet cells. The size of the absorptive cell localizations and their lack of association with any discernable cell structure led to a re-evaluation of the oxalate precipitation technique for the maintenance of intracellular calcium and to the development of a new preparative method, independent of any exogenous precipitating agent. The new method is based on freeze drying and is shown to preserve the in vivo distribution of Na, K, P and Ca. The calcium distribution using this technique was found to involve higher concentrations of cytoplasmic calcium, but considerably fewer and much smaller localizations than found with the oxalate technique.