We describe an assay for ethylene glycol in serum, which is based on the Hantzsch condensation reaction. Ethylene glycol is dehydrogenated by sodium periodate to form two molecules of formaldehyde, which is then combined with ammonium ion and acetylacetone to form a fluorescent diacetyllutidine derivative. The detection limit of this method is 50 mg/L. The procedure is sensitive, requires only 100 micro L of serum, and the standard curve is linear over a working range of 50 to 250 mg/L. Average analytical recovery ranged from 99 to 102% and within-run precision studies showed CVs of < 5%. Reagents are easily prepared and available.