LOCATION-DEPENDENT ARTIFACT FOR NO MEASUREMENT USING MULTIWELL PLATES

The Griess reaction is widely used to measure the cellular production of NO by detecting the supernatant levels of nitrite. Ordinarily, back ground levels of nitrite in the media are subtracted from the levels o f nitrite produced by the cells by preparing a ''blank'' during the de termination of the standard curve. Although this method is adequate fo r most experimental conditions, it cannot be used when cell supernatan ts are collected from multiwell dishes, particularly when low amounts of NO are produced and when long incubation periods are required to in duce NO production. Our data show that a highly variable level of nitr ite is found in the absence of cells in the media from wells at the ed ges of the 96-well plate while media from interior wells shows no dete ctable nitrite accumulation. The most likely source of this noncellula r NO is from nitric oxides (NOx) found in the ambient air and reductio n of air exchange or regulation of the gaseous environment eliminates this ''border effect.''