We present TrAQ, a new MATLAB-based two-dimensional tracking software for Open Field video analysis of unmarked single animal, featuring minimum user intervention. We developed TrAQ with the purpose to automatically count the in-plane rotations, an important parameter in the 6-hydroxydopamine hemiparkinsonian rat model and in many rodent models of neurodegenerative diseases, a very time-consuming manual task for highly trained human operators. In addition, TrAQ allows automatic recognition of the animal within a user defined arena providing a quantitative measurement of the body centroid and the two extremities positions. A full range of quantitative kinematic behavioral parameters are automatically calculated, and the optional shape erosion feature adds usage flexibility. TrAQ, free and non-species-specific application, was quantitively tested with rodents and on a qualitative basis with zebrafish, and invertebrate animal models. Quantitative results were successfully validated against a commercial software (for tracking) and manual annotation (for rotations in an hemiparkinsonian rat model). This is a widely used model in preclinical research to study postural instability and motor asymmetry. TrAQ allows the characterization of motor asymmetry using non-invasive tools, thus appreciating the spontaneous Open Field behaviour of unmarked single animal, with minimum user intervention.