The concept of contact inhibition of locomotion (CIL) describes the ability of a cell to change the direction of its movement after contact with another cell. It has been shown to be responsible for physiological and developmental processes such as wound healing, macrophage dispersion and neural crest cell migration; whereas its loss facilitates cancer cell invasion and metastatic dissemination. Different assays have been developed to analyze CIL in tissue culture models. However, these methods have several caveats. Collisions happen at low frequency between freely migrating cells and the orientation of the cells at the time of contact is not predictable. Moreover, the computational analysis required by these assays is often complicated and it retains a certain degree of discretion. Here, we show that confinement of neural crest cell migration on a single dimension by using a micropatterned substrate allows standardized and predictable cell-cell collision. CIL can thus easily be quantified by direct measurement of simple cellular parameters such as the distance between nuclei after collision. We tested some of the signaling pathways previously identified as involved in CIL, such as small GTPases and non-canonical Wnt signaling, using this new method for CIL analysis. The restricted directionality of migration of cells in lines is a powerful strategy to obtain higher predictability and higher efficiency of the CIL response upon cell-cell collisions.