We used a surfactant to fabricate a microfluidic paper-based analytical device (mu PAD) by selective modification of hydrophobic filter paper. Hydrophobic tails of the surfactant penetrate and remain in the hydrophobic layer of filter cellulose while hydrophilic heads extend out of the hydrophobic layer, forming a hydrophilic layer on the hydrophobic cellulose to direct flow of aqueous solutions. A flower-shaped mu PAD was fabricated by the presented method and was applied to the protein assay in human urine.