Background: Waiting temperature before centrifugation and anticoagulants used, markedly effect total homocysteine concentrations. The aim of this study was to investigate the effect of different anticoagulants and temperature on plasma homocysteine levels. Methods: We studied total homocysteine concentrations in 23 healthy subjects. Blood was drawn in K(3)EDTA, sodium citrate- or sodium fluoride-containing tubes, and kept at 0 degreesC or 22 degreesC for 3 h. Total homocysteine measurements were performed with fluorescence polarization immunoassay (FPIA) method. We compared all results with baseline EDTA values (samples put on crushed ice and centrifuged immediately) recommended in literature for reference handling. Results: At 22 degreesC, the tubes: containing sodium citrate and sodium fluoride showed significantly higher total homocysteine concentrations than their respective baseline values (p = 0.000). However, sodium fluoride tubes were not significantly different than baseline EDTA levels. Waiting 3 h at 0 degreesC did not effect sodium citrate and EDTA plasma total homocysteine concentrations when compared to baseline EDTA, but sodium fluoride-containing plasma levels were significantly decreased( p = 0.000). Conclusions: According to our results, the most available and practical temperature and anticoagulant for total homocysteine determination is sodium fluoride at room temperature up to 3 h. (C) 2001 Elsevier Science B.V. All rights reserved.