Transcranial magnetic stimulation (TMS) is a non-invasive method to investigate motor pathways and to create a map of the somatotopical organization of the motor cortex: ordinary mapping procedures requires a focal brain stimulation over different spots of the scalp and electromyographic (EMG) recording from a muscle. Finding an appropriate and a valid visual representation of collected data is a crucial step in research and clinical field to allow a relatively fast, intra- and inter-patient comparison of motor cortex mapping. Aim of this study was to develop and to validate a method to map cortical representation of an intrinsic hand muscle (abductor digiti minimi, ADM) using a two-dimensional spline interpolation of EMG peak amplitudes obtained with TMS. The interpolated model will result in a graphical colour-scaled representation of the motor cortex for the investigated muscle; fitted model was finally validated by comparing derived parameters with those directly measured to ensure the strength and reliability of the model. Ten healthy volunteers (mean age+/-S.D.: 35.3+/-4.7 years, 4 males and 6 females) were enrolled in the study. Transcranial stimulation was performed by placing a figure-of-eight coil over a predefined grid on the scalp of the subject. EMG responses were recorded from the right abductor digiti minimi (ADM): averaged EMG peak amplitudes obtained at each node were then used to perform spline interpolation and to derive other parameters like center of gravity (CoG). Arithmetical mean of all resting motor threshold at the hotspot was 50.6+/-3.4% of the maximal stimulator output. Average amplitude at the hotspot was 1.72+/-0.80 mV and its coordinates, expressed as median, were x=4.5 cm and y=0.0 cm. Mean CoG was located at x=4.86+/-0.57 cm and y=0.35+/-0.10 cm. Mean interpolated peak coordinates for ADM were xf=4.86+/-0.58 cm and yf=0.36+/-0.12 cm, while mean fitted peak amplitude was 0.87+/-0.47 mV. Results suggest how it is possible to map the primary motor cortex using two-dimensional spline interpolation of peak-to-peak amplitudes obtained by single pulse TMS delivered on several scalp positions, which will result in a smooth, easy to read, colour-scaled map. However, like other visual representation modalities, the interpolation should become complementary to traditional methods and not a substitute of a precise and accurate cortical motor mapping.