Folate content in berries and the effect of cultivar on folate content in strawberries were investigated using a modified radioprotein-binding assay (RPBA) combined with solid phase extraction (SPE). With an SPE step prior to quantification the matrix effects in the RPBA assay were minimised. HPLC quantification was used to compare and characterise individual folate forms. High correlation (r = 0.9796) was shown and the mean difference between the RPBA and HPLC method was 6.7 4.8 mug/100 g. The predominating folate form was 6S-5-CH3-H-4-folate in all tested berries (>95%). Therefore, this biologically active folate form (6S-5-CH3-H(4)folate) must be chosen as calibrant because the RPBA response of the synthetic equimolar mixture of R- and S-diastereomers (6R,S-5-CH3-H(4)folate) was considerably higher, about 30%. The folate content in nine selected Swedish berries varied from 11 to 96 mug/100 g (fresh weight) and the highest folate content was found in rose hips (96 mug/100 g) and strawberries (74 mug/100 g). Moderate folate sources were raspberry (46 mug/100 g) and sea buckthorn (39 mug/100 g), whereas all other tested berries contained about 20 mug of folate/100 g or less. In strawberries, folate content varied significantly among eight different cultivars from 73 mug/100 g (Lina) to 99 mug/100 g (Melody).