High-resolution 3D printed microfluidic immunoaffinity solid-phase extraction columns

Solid-phase extraction is essential for sample preparation and biomarker enrichment in complex biological and environmental matrices, yet conventional packed-bed SPE columns suffer from heterogeneous particle packing, poor mass transfer, and limited geometric control that constrain selectivity and scalability. Alternative formats such as polymer or silica monoliths and micromachined pillar arrays offer improved flow and mass transfer but remain limited in broad commercial implementation. Here, we demonstrate rapid fabrication of immunoaffinity solid-phase extraction columns using high-resolution stereolithographic 3D printing. Triply periodic minimal surface structures were printed at a rate exceeding 20 columns per hour using a polyethylene glycol diacrylate based resin. Scanning electron microscopy confirmed well-defined triply periodic minimal surface geometries with tunable pore sizes from 10 to as small as 2.5 μm. The epoxide end of glycidyl methacrylate doped into the resin enabled covalent immobilization of anti-red fluorescent protein antibodies to generate an affinity stationary phase. Affinity extraction performance was evaluated by laser-induced fluorescence monitoring of red fluorescent protein, demonstrating efficient retention and preconcentration after loading and washing, with a distinct peak upon elution. Selectivity was then demonstrated by extracting red fluorescent protein from a multicomponent mixture. These results establish high-resolution 3D printing as a scalable platform for fabricating customizable immunoaffinity columns with precise control over pore architecture and surface chemistry, enabling miniaturized sample preparation for bioanalytical applications.