The capability of modifying and patterning the surface of polymer and composite materials is of high significance for various biomedical and electronics applications. For example, the use of femtosecond (fs) laser ablation for micropatterning electrospun nanofiber scaffolds can be successfully employed to fabricate complex polymeric biomedical devices, including scaffolds. Here we investigated fs-laser ablation as a flexible and convenient method for micropatterning polyamide (PA6) electrospun nanofibers that were modified with molybdenum disulfide (MoS2). We studied the influence of the laser pulse energy and scanning speed on the topography of electrospun composite nanofibers, as well as the irradiated areas via scanning electron microscopy and spectroscopic techniques. The results showed that using the optimal fs-laser parameters, micropores were formed on the electrospun nanofibrous membranes with size scale control, while the nature of the nanofibers was preserved. MoS2-modified PA6 nanofibrous membranes showed good photoluminescence properties, even after fs-laser microstructuring. The results presented here demonstrated potential application in optoelectronic devices. In addition, the application of this technique has a great deal of potential in the biomedical field, such as in tissue engineering.