AR protein interrogation in CTCs from patients with metastatic castration-resistant prostate cancer using ImageStreamX. A. Flow chart illustrating the protocol for preparing CTCs for analysis using ImageStreamX. B. Representative dot plot showing area (cell size) and AR intensity (expression) for a single patient. Two CTCs have been selected to show corresponding images acquired using ImageStreamX technology, which demonstrate AR-Low (top panel) or AR-high (bottom panel) expression. C. Representative histogram showing nuclear or non-nuclear AR for all CTCs from a single patient. Nuclear AR is calculated by the similarity of AR staining and FxCycle Violet staining, and defined as Similarity Index >1. Two CTCs have been selected to show corresponding images acquired using ImageStreamX technology.