Three-Dimensional Paper-Based Microfluidic Analytical Devices Integrated with a Plasma Separation Membrane for the Detection of Biomarkers in Whole Blood

Paper-based microfluidic analytical devices (mu PADs) have recently attracted attention as a point-of-care test kit because of their low cost and nonrequirement for external forces. To directly detect biomarkers in whole blood, however, they need to be assembled with a filter such as a plasma separation membrane (PSM) because the color of the blood cells interferes with the colorimetric assay. However, this assembly process is rather complicated and cumbersome, and the fluid does not uniformly move to the detection zone when the adhesion between the paper and PSM is not perfect. In this study, we report a simple three-dimensional (3D) printing method for fabricating PSM-integrated 3D-mu PADs made of plastics without the need for additional assembly. In detail, PSM was coated with parylene C to prevent its dissolution from organic solvent during 3D printing. Then, the coated PSM was superimposed on the paper. Detection zones and a reservoir were printed on the paper and PSM via liquid photopolymerization, using a digital light processing printer. The limit of detection of the PSM-integrated 3D-mu PADs for glucose in whole blood was 0.3 mM, and these devices demonstrated clinically relevant performance on diabetes patient blood samples. Our 3D-mu PADs can also simultaneously detect multiple metabolic disease markers including glucose, cholesterol, and triglycerides in whole blood. Our results suggest that our printing method is useful for fabricating 3D-mu PADs integrated with PSM for the direct detection of biomarkers in whole blood.