A study was made of the conditions necessary for the quantitative assay of the erythrocyte and plasma cholinesterases of the rat by the Warburg manometric method. Butyrylcholine, acetyl-beta-methylcholine, and acetylcholine were tested as substrates, but only acetylcholine proved satisfactory. The level of the red cell cholinesterase did not change with age of the rat. The erythrocytes were found to contain an esterase which hydrolyzes butyrylcholine. In confirmation of other workers, it was found that the plasma cholinesterase levels of female rats increase with age and are higher than those of male rats of all ages and that the plasma of male animals contains an approximately equal mixture of true and pseudocholinesterase whereas that of female rats contains mainly pseudocholinesterase.