Protocol for establishing primary human lung organoid-derived air-liquid interface cultures from cryopreserved human lung tissue

  • STAR Protoc. 2023 Dec 15;4(4):102735. doi: 10.1016/j.xpro.2023.102735.
Diana Cadena Castaneda  1 ,  Sonia Jangra  2 ,  Marina Yurieva  1 ,  Jan Martinek  1 ,  Megan Callender  1 ,  Matthew Coxe  1 ,  Angela Choi  2 ,  Juan García-Bernalt Diego  2 ,  Te-Chia Wu  1 ,  Florentina Marches  1 ,  Damien Chaussabel  1 ,  Adolfo García-Sastre  3 ,  Michael Schotsaert  2 ,  Adam Williams  4 ,  Karolina Palucka  5
Affiliations
  • 1. The Jackson Laboratory for Genomic Medicine, Farmington, CT 06032, USA.
  • 2. Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Global Health and Emerging Pathogens Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA.
  • 3. Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Global Health and Emerging Pathogens Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Department of Medicine, Division of Infectious Diseases, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; The Tisch Cancer Institute, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA; Department of Pathology, Molecular and Cell-Based Medicine, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA.
  • 4. The Jackson Laboratory for Genomic Medicine, Farmington, CT 06032, USA. Electronic address: [email protected].
  • 5. The Jackson Laboratory for Genomic Medicine, Farmington, CT 06032, USA. Electronic address: [email protected].
PMID: 37991921 DOI: 10.1016/j.xpro.2023.102735
Abstract

Primary human lung organoid-derived air-liquid interface (ALI) cultures serve as a physiologically relevant model to study human airway epithelium in vitro. Here, we present a protocol for establishing these cultures from cryopreserved human lung tissue. We describe steps for lung tissue cryostorage, tissue dissociation, lung epithelial Organoid generation, and ALI culture differentiation. We also include quality control steps and technical readouts for monitoring virus response. This protocol demonstrates severe acute respiratory syndrome coronavirus 2 Infection in these cultures as an example of their utility. For complete details on the use and execution of this protocol, please refer to Diana Cadena Castaneda et al. (2023).1.

Keywords
Immunology; Microscopy; Organoids.