Protocol for generating airway organoids from 2D air liquid interface-differentiated nasal epithelia for use in a functional CFTR assay.

We present a protocol to generate organoids from air-liquid-interface (ALI)-differentiated nasal epithelia. We detail their application as cystic fibrosis (CF) disease model in the cystic fibrosis transmembrane conductance regulator (CFTR)-dependent forskolin-induced swelling (FIS) assay. We describe steps for isolation, expansion and cryostorage of nasal brushing-derived basal progenitor cells, and their differentiation in ALI cultures. Furthermore, we detail the conversion of differentiated epithelial fragments into organoids of healthy controls and CF subjects for validating CFTR function and modulator responses. For complete details on the use and execution of this protocol, please refer to Amatngalim et al. ¹ .
Competing Interests: Declaration of interests J.M.B. has a patent granted (10006904) related to CFTR function measurements in intestinal organoids and received personal fees from HUB/Royal Dutch academy of sciences, during the conduct of the study. He received nonfinancial support from Vertex Pharmaceuticals and personal fees and nonfinancial support from Proteostasis Therapeutics, outside the submitted work.
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