The Batch-Resourcing Angiogenesis Tool (BRAT) to enable high-throughput microscopy screening of microvascular networks

Vessel forming assays are a valuable technology to evaluate the vasculogenic and angiogenic potential of different cell types, matrix proteins, and soluble factors. Recent advances in high-content microscopy allow for vascular morphogenesis assays to be captured in real-time and in high-throughput formats. Unfortunately, existing microvascular network (MVN) quantification algorithms are either inaccurate, not user-friendly, or manually analyse one image at a time, unfavourable to high-throughput screening. This manuscript introduces the Batch-Resourcing Angiogenesis Tool (BRAT), an open-source computer software which efficiently segments, skeletonizes, and analyses large batches of vascular network images with high accuracy. Benchmarked across diverse clinical and cultured MVN images, BRAT is the most sensitive vascular network image analysis tool (94.5%), exhibiting leading accuracy (93.3%). BRAT's multi-threaded processing automatically analyses 886 microscopy images at a speed of 0.17 s/image on a performance computer (2:29 min) or 2.31 s/image on a laptop (34:04). This is 10-to-100 fold more time-efficient than existing software, which require 12-16 s of direct user input per image. BRAT successfully compares diverse microvascular cell types cultured in 2D and 3D biomaterials. BRAT represents a powerful approach for the accurate and high-throughput screening of vessel forming assays for disease models, regenerative medicines, and therapeutic testing. BRAT is avaliable to download at:https://github.com/BMSE-UQ/BRAT-Vascular-Image-Tool.