3D-carbon nanomaterials have proven to be high-performance transducers in electrochemical sensors but their integration into miniaturized devices is challenging. Herein, we develop printable freestanding laser-induced carbon nanofibers (f-LCNFs) with outstanding analytical performance that furthermore can easily allow such miniaturization through a paper-based microfluidic strategy. The f-LCNF electrodes were generated from electrospun polyimide nanofibers and one-step laser carbonization. A three-electrode system made of f-LCNFs exhibited a limit of detection (LOD) as low as 1 nM (S/N= 8) for anodic stripping analysis of silver ions, exhibiting the peak at ca. 100 mV vs f-LCNFs RE, without the need of stirring. The as-described system was implemented in miniaturized devices via wax-based printing, in which their electroanalytical performance was characterized for both outer- and inner-sphere redox markers and then applied to the detection of dopamine (the peak appeared at ca. 200 mV vs f-LCNFs RE) with a remarkable LOD of 55 pM. When modified with Nafion, the f-LCNFs were highly selective to dopamine even against high concentrations of uric and ascorbic acids. Especially the integration into closed microfluidic systems highlights the strength 3D porous structures provides excellent analytical performance paving the way for their translation to affordable lab-on-a-chip devices where mass-production capability, unsophisticated fabrication techniques, transfer-free, and customized electrode designs can be realized.