BEADS: Un flujo de trabajo interactivo y semiautomatizado para ensayos de angiogénesis en microesferas 3D que permite...

3D fibrin bead angiogenesis assays are widely used to study endothelial sprouting in vitro, yet current analytical approaches are either time-consuming or poorly adaptable to complex imaging conditions, limiting quantitative assessment of co-cultures, spatial interactions, and nearest-neighbor-dependent angiogenic behavior. In this study, we developed a semiautomated user-interactive image analysis pipeline, Bead-based Endothelial Angiogenesis Data Suite (BEADS), to provide standardized quantitative bead-centric metrics of sprouting, migration, and spatial orientation in 3D fibrin angiogenesis assays. BEADS integrates automated bead detection with manual correction, followed by guided sprout and migratory-cell annotation across multi-channel image z-stacks. Novel analytical capabilities include co-culture designation, nearest-neighbor pairing, and circular statistics for sprout-directionality quantification. Performance was evaluated in assays using co-cultured male and female human pulmonary microvascular endothelial cell (HPMEC)-coated beads. BEADS reduced hands-on analysis time approximately sevenfold compared with manual tracing while preserving sprout-length accuracy against manual ground truth. BEADS provides a standardized, extensible platform for microvascular image analysis, supporting co-culture experimentation, spatial endothelial-interaction metrics, migratory-cell quantification, and high-throughput adaptation. This semiautomated workflow enables quantitative microvascular research by integrating computational precision with endothelial behavior and is broadly applicable to angiogenesis assays that incorporate co-cultures, perturbations, or multi-label experimental designs.