The 3D tracking of micro-objects, based on digital holography, is proposed through the analysis of the complex wavefront of the light scattered by the micro-samples. Exploiting the advantages of the off-axis full-field holographic interferometry, the tracking of multiple objects is achieved by a direct wavefront analysis at the focal plane overcoming the limitation of the conventional back focal plane interferometry in which only one object at a time can be tracked. Furthermore, the method proposed and demonstrated here is a step forward with respect to other holographic tracking tools. The approach is tested in two experiments, the first investigates the Brownian motion of particles trapped by holographic optical tweezers, while the second relates to the cell motility in a 3D collagen matrix, thus showing its usefulness for lab-on-chip systems in typical bioassay testing.