Ethylene glycol ingestion, accidental or intentional, can be a life-threatening emergency. Assays are not available from most clinical laboratories, and, thus, results often require many hours or days to obtain. Enzymatic assays, adaptable to automated chemistry analyzers, have been evaluated, but they have been plagued by analytic problems. With an alternative method of data analysis applied to an existing enzymatic assay, a modified assay was developed and validated on 2 different automated chemistry systems. Compared with a previously validated method based on gas chromatography with flame ionization detection, the modified enzymatic assay showed excellent agreement on patient samples (y = 1.0227x -1.24; r(2) = 0.9725), with a large analytic measuring range (2.5-300 mg/dL [0.4-48.4 mmol/L]). Interferences from propylene glycol, various butanediols, and other related compounds were almost entirely eliminated; when present, they generated error flags rather than falsely elevated ethylene glycol results. This modified assay should make it possible for more clinical laboratories to offer ethylene glycol measurements.