We have developed an automated chromogenic peptide substrate assay for factor XII (FXIIcs) on a Cobas Mira S Plus clinical chemistry analyser using a new commercially available kit. This was used to determine factor XII (FXII) levels in plasma samples from 320 blood donors, 206 patients with a history of venous thrombosis and 74 lupus anticoagulant positive (LA+) patients. Results were compared with those obtained in a clotting assay for FXII (FXIIct) and an immunochemical assay (FXIIag). A satisfactory correlation coefficient of 0.92 and a regression line equation of y = 7.898 + 0.871x was obtained between FXIIcs and FXIIct in the 320 blood donors. Levels of FXII below the calculated normal range were found in nine blood donors (2.8%) and 16 venous thrombosis patients (7.8%). The blood donors and patients with venous thrombosis with low FXIIcs values had FXII levels below our lower limits of normal for both FXIIct and FXIIag; all were lupus anticoagulant negative. When FXII levels were determined in the 74 LA+ patients, 27 (36.5%) gave markedly lower FXII values in the FXIIct when compared with the FXIIcs. FXIIag levels corresponded with FXIIcs. The automated FXIIcs assay is therefore lupus anticoagulant insensitive and allows us to measure FXII levels accurately and routinely in large numbers of patient samples. Blood Coag Fibrinol 9:183–187 × 1998 Lippincott-Raven Publishers.