A direct photometric method for assaying neurotoxic esterase (NTE) activity of chicken brain microsomal preparation has been developed using 4-nitrophenyl esters as substrates. Paired samples of the microsomal preparation were preincubated for 20 min. with paraoxon plus either (a) buffer or (b) mipafox before addition of substrate. The initial rate of NTE activity was directly recorded at 410 nm by matching the content of tube (a) against tube (b) after addition of the substrate to both of them. The 4-nitrophenyl esters of propionic, butyric, valeric, lauric, capric and caproic acids were tested as substrates. Results indicated that 4-nitrophenyl valerate and caporate, respectively, are the most hydrolyzable substrates for NTE with this method; it also enables detained kinetic studies of NTE to be made. The Michaelis constant (Km) for the hydrolysis of 4-nitrophenyl valerate by NTE was found to be 0.0000555M.