A procedure is described for the fully automated quantitative determination of platelet-activating factor (PAF) using liquid chromatography/fast atom bombardment mass spectrometry (LC/FAB-MS) with two types of column-switching techniques. Various parameters in LC/FAB-MS were optimized, and the use of a microbore column allowed highly sensitive detection of PAF. A column-switching system was incorporated to minimize band broadening and to increase the permissible injection volume. Analysis of authentic PAF indicated that the limit of quantitation was at the low picogram level (about 50 pg/ml). Another system using both a strong cation-exchange column and an ODS column in tandem was developed for the analysis of PAF in blood samples. These methods were validated with standard samples and applied to the determination of PAF in human polymorphonuclear neutrophils stimulated by addition of a calcium ionophore and in human blood.