Angiogenesis is a critical, fine-tuned, multi-staged biological process. Tip-stalk cell selection and shuffling are the building blocks of sprouting angiogenesis. Accumulated evidences show that tip-stalk cell selection and shuffling are regulated by a variety of physical, chemical and biological factors, especially the interaction among multiple genes, their products and environments. The classic Notch-VEGFR, Slit-Robo, ECM-binding integrin, semaphorin and CCN family play important roles in tip-stalk cell selection and shuffling. In this review, we outline the progress and prospect in the mechanism and the roles of the various molecules and related signaling pathways in endothelial tip-stalk cell selection and shuffling. In the future, the regulators of tip-stalk cell selection and shuffling would be the potential markers and targets for angiogenesis.