Oxidative stress has been implicated in a wide variety of disease processes. One method to quantify oxidative injury is to measure lipid peroxidation. Most methods to do this are fraught with problems particularly when utilized to assess oxidative stress in vivo. On the other hand, quantification of a group of prostaglandin F(2)-like compounds, termed the F(2)-isoprostanes (F(2)-IsoPs) provides an accurate assessment of oxidative stress both in vitro and in vivo and has come to be regarded as the gold standard to quantify lipid peroxidation. This unit describes methods to assess lipid peroxidation associated with oxidant injury in vivo by quantifying concentrations of either esterified or free F(2)-IsoPs in biological fluids and tissues. The techniques employed for the analysis of these compounds from biological sources that are detailed herein utilize mass spectrometric approaches. Measurement of F(2)-IsoPs represents an important advance in the ability to assess the role of oxidative stress in human disease.