The measurement of low levels of cholinesterase or acetylcholinesterase by the Ellman method requires correction for a non-enzymatic increase in absorption at 412 mμ that is due both to non-enzymatic hydrolysis of the acetylthiocholine substrate and to modification of the colour reagent. The rate of increase in absorption is dependent on temperature and pH. Addition of an acidic solution of lysivane to the assay solution for selective measurement of amniotic fluid acetylcholinesterase gives rise to a shift in pH; the use of methanol is suggested as an easier method of dissolving the inhibitor and does not affect the pH of the assay, obviating any need to redetermine the background absorption. There is, however, no improvement in ability of the method to predict pregnancies associated with neural-tube defects.