A cDNA clone was obtained from Arabidopsis thalianathat encodes a protein containing 92 amino acid residues with high sequence identity (57%) to bovine acyl-CoA-binding protein (ACBP). The coding sequence of this clone was expressed in Escherichia coliand the gene product (10.4 kDa) was purified. The recombinant A. thalianaACBP (rAthACBP) was shown to bind acyl-CoA esters and protect acyl-CoAs from degradation by microsomal acyl-hydrolases. Antibodies that were raised to rAthACBP recognized the native ArabidopsisACBP and also cross-reacted with a number of other plant ACBPs, including rapeseed ( Brassica napus) ACBP. The pattern of expression and level of the gene product were examined in various tissues of Arabidopsisand Brassicausing Western blotting. A. thalianatissues contained between 3 and 143 μg AthACBP g −1FW depending on the tissue (0.4 to 14 nmol g −1FW). Developing B. napusseeds underwent a 12-fold increase in ACBP levels during seed maturation (20 to 250 μg ACBP g −1FW); the highest concentration occurring near the peak of triacylglycerol accumulation (26 nmol g −1FW).