The novel method for biogenic component determination in liquid fuels by direct measurement of the C-14 activity concentration via liquid scintillation counting (LSC) technique has been used in few laboratories worldwide. This paper presents results of the development of method for bio-component quantification in fuel mixtures in the Nuclear Physics Laboratory Novi Sad, Serbia, as well as their comparison to established methodology in literature with intent to define optimal parameters and measurement conditions for 14C determination in biofuels. Therefore, the purpose of the paper is to give comprehensive overview of optimal parameters such as scintillation cocktail and vial selection, fuel/cocktail ratio and best calibration procedure of liquid scintillation counter Quantulus 1220 (TM), with practical information for laboratories that intend to develop this technique. Biodiesel produced from two different feedstock materials was mixed with two commercial fossil matrices to produce fuel blends containing 1-100% FAME (Fatty Acid Methyl Esters), which were analyzed for calibration purposes. Analytical study of relation between the detected released energy and quench level led to establishment of calibration curves specific to the fuel types produced in this moment on the market in Serbia. (C) 2016 Elsevier Ltd. All rights reserved.