Recent interest in the folate status of diverse populations as a serious clinical and public health issue has been associated with substantial demands for blood folate analyses. Folate analysis by microbiological assays fulfills two important requirements for such screening by providing an acceptable detection limit and specificity at a low reagent cost. Dried blood spots (DBS) on special filter paper have been notably cost-effective as a blood sample collection device in epidemiologic field studies, allowing fingerstick peripheral blood sampling and ease of specimen transport and storage. We recently combined the advantages of a microbiological assay and DBS in an assay for erythrocyte folate (1). We now describe a dried-serum spot (DSS) assay for serum folate (SF). Filter-paper cards, each with 15 preprinted circles, were used (Type 903; Schleicher & Schuell). Individual fresh serum or EDTA-plasma samples from clinic patients or healthy donor volunteers were used throughout. DSS were prepared by pipetting 50–150 μL of serum onto the paper and drying the papers on a rack for at least 5 h at room temperature before storage at −80 °C in resealable plastic bags with desiccant sachets (1). Some paper was preimpregnated with ascorbic acid in an effort to extend DSS folate stability. These 15 × 10-cm paper cards were immersed flat in 10 mL of 10 g/L ascorbic acid for 1 min, followed by air-drying on sheets of aluminum foil. Whole DSS folates were analyzed after the entire serum spot (50 μL) was cut out using scissors. The spots were cut in half and placed in the base of a borosilicate tube (12 × 75 mm), covered with 2 mL of 5 g/L sodium ascorbate containing 0.5 mL/L Tween 80, mixed on a vortex-mixer, and sonicated for 15 min (Deacon Ultrasonics Ltd). The DSS eluate was considered to represent a 1:40 …