4.2. EBUS-TBNA

We performed all clinical procedures with patients under deep sedation using a flexible ultrasound bronchoscope (CP-EBUS-XBF-UC260F, Olympus, Tokyo, Japan) and a 21G Vizashot needle. At least three transbronchial needle aspirations were taken at each suspected tumor or nodal station in accordance with diagnostic and staging standards. The initial samples were processed in a standard institutional fashion previously described [63], with preparation of slides for rapid on-site cytological interpretation, as well as collecting material to build a cell-block for delayed pathological evaluation. If rapid on-site cytology reported a high likelihood of tumor from a specified location, a fine needle aspirate (FNA) from the same location was collected and expelled into ice-cold culture media (for PDX generation) or RNAlater (for expression analysis). In addition to the needle aspiration, study participants underwent mucosal brushings of the right proximal main-stem bronchus using a standard, disposable cytology brush. Two brushings were obtained from each subject and stored in RNAlater solution (ThermoFisher).