We have designed a 2-spinnerette device that can directly electrospin nanofiber scaffolds containing a gradient in composition that can be used to engineer interfacial tissues such as ligament and tendon. Two types of nanofibers are simultaneously electrospun in an overlapping pattern to create a nonwoven mat of nanofibers containing a composition gradient. The approach is an advance over previous methods due to its versatility - gradients can be formed from any materials that can be electrospun. A dye was used to characterize the 2-spinnerette approach and applicability to tissue engineering was demonstrated by fabricating nanofibers with gradients in amorphous calcium phosphate nanoparticles (nACP). Adhesion and proliferation of osteogenic cells (MC3T3-E1 murine pre-osteoblasts) on gradients was enhanced on the regions of the gradients that contained higher nACP content yielding a graded osteoblast response. Since increases in soluble calcium and phosphate ions stimulate osteoblast function, we measured their release and observed significant release from nanofibers containing nACP. The nanofiber-nACP gradients fabricated herein can be applied to generate tissues with osteoblast gradients such as ligaments or tendons. In conclusion, these results introduce a versatile approach for fabricating nanofiber gradients that can have application for engineering graded tissues. © The Author(s) 2012.