Detailed protocol for a corneal thermal cauterization-based (lymph-)angiogenesis assay in mice

Angiogenesis and lymphangiogenesis, the formation of new blood or lymphatic vessels, respec-tively, from preexisting vasculature is essential during embryonic development, but also occurs during tissue repair and in pathological conditions (cancer; ocular disease; ischemic, infectious and inflammatory disorders), which are all characterized to a certain extent by inflammatory con-ditions. Hence, a rapid, inexpensive, feasible / technically easy, reliable assay of inflammation -induced (lymph-)angiogenesis is highly valuable. In this context, the corneal thermal cauterization assay in mice is a simple, low-cost, reproducible, insightful and labor-saving assay to gauge the role of inflammation in angiogenesis and lymphangiogenesis. However, to the best of our knowl-edge, there is no standardized protocol to perform this assay. Here, we provide a step-by-step description of the model's procedures, which include:center dot The thermal cauterization of the corneas,center dot Enucleation and dissection of the corneas,center dot Subsequent immunofluorescence staining of the neovasculature, and morphometric analysis.We also discuss ethical considerations and aspects related to animal welfare guidelines. Alto-gether, this paper will help to increase the reproducibility of the corneal thermal cauterization model and facilitate its use for angiogenesis and lymphangiogenesis research.