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To facilitate increased regeneration speed and functional recovery, the ideal conduit should provide biochemically relevant signals and physical guidance cues, thus playing an active role in regeneration. To that end, laminin and laminin\u2013polycaprolactone (PCL) blend nanofibers were fabricated to mimic peripheral nerve basement membrane. <jats:italic>In vitro<\/jats:italic> assays established 10% (wt) laminin content is sufficient to retain neurite\u2010promoting effects of laminin. In addition, modified collector plate design to introduce an insulating gap enabled the fabrication of aligned nanofibers. The effects of laminin content and fiber orientation were evaluated in rat tibial nerve defect model. The lumens of conduits were filled with nanofiber meshes of varying laminin content and alignment to assess changes in motor and sensory recovery. Retrograde nerve conduction speed at 6 weeks was significantly faster in animals receiving aligned nanofiber conduits than in those receiving random nanofiber conduits. Animals receiving nanofiber\u2010filled conduits showed some conduction in both anterograde and retrograde directions, whereas in animals receiving hollow conduits, no impulse conduction was detected. Aligned PCL nanofibers significantly improved motor function; aligned laminin blend nanofibers yielded the best sensory function recovery. In both cases, nanofiber\u2010filled conduits resulted in better functional recovery than hollow conduits. These studies provide a firm foundation for the use of natural\u2013synthetic blend electrospun nanofibers to enhance existing hollow nerve guidance conduits. \u00a9 2011 Wiley Periodicals, Inc. 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