Lyme disease is the most important vector-borne disease in the Northern hemisphere and represents a major public health challenge. The disease diagnosis relies mainly on clinical practice that can be coupled to serological or molecular diagnostics. However the relatively low sensitivity and specificity of the available Lyme diagnostics indicate the urgent need to identify additional highly antigenic borrelial proteins or to develop new strategies of diagnostic. As the skin constitutes a key interface where the pathogens can persist and multiply, we investigated proteomics on skin samples to detect Borrelia proteins directly in cutaneous biopsies in a robust and specific way. We first set up a discovery Ge-LC-MS/MS approach on a murine model infected by B. burgdorferi sensu stricto. This dataset contains all the results concerning this approach.