Display options
Cite
Vonk AM, van Mourik P, Ramalho AS, et al. Protocol for Application, Standardization and Validation of the Forskolin-Induced Swelling Assay in Cystic Fibrosis Human Colon Organoids. STAR Protoc. 2020;1(1):100019doi: 10.1016/j.xpro.2020.100019.
Vonk, A. M., van Mourik, P., Ramalho, A. S., Silva, I. A. L., Statia, M., Kruisselbrink, E., Suen, S. W. F., Dekkers, J. F., Vleggaar, F. P., Houwen, R. H. J., Mullenders, J., Boj, S. F., Vries, R., Amaral, M. D., de Boeck, K., van der Ent, C. K., & Beekman, J. M. (2020). Protocol for Application, Standardization and Validation of the Forskolin-Induced Swelling Assay in Cystic Fibrosis Human Colon Organoids. STAR protocols, 1(1), 100019. https://doi.org/10.1016/j.xpro.2020.100019
Vonk, Annelotte M, et al. "Protocol for Application, Standardization and Validation of the Forskolin-Induced Swelling Assay in Cystic Fibrosis Human Colon Organoids." STAR protocols vol. 1,1 (2020): 100019. doi: https://doi.org/10.1016/j.xpro.2020.100019
Vonk AM, van Mourik P, Ramalho AS, Silva IAL, Statia M, Kruisselbrink E, Suen SWF, Dekkers JF, Vleggaar FP, Houwen RHJ, Mullenders J, Boj SF, Vries R, Amaral MD, de Boeck K, van der Ent CK, Beekman JM. Protocol for Application, Standardization and Validation of the Forskolin-Induced Swelling Assay in Cystic Fibrosis Human Colon Organoids. STAR Protoc. 2020 Jun 03;1(1):100019. doi: 10.1016/j.xpro.2020.100019. eCollection 2020 Jun 19. PMID: 33111074; PMCID: PMC7580120.
Copy Download .nbib Format: NLM
Share it on

STAR Protoc. 2020 Jun 03;1(1):100019. doi: 10.1016/j.xpro.2020.100019. eCollection 2020 Jun 19.

Protocol for Application, Standardization and Validation of the Forskolin-Induced Swelling Assay in Cystic Fibrosis Human Colon Organoids.

STAR protocols

Annelotte M Vonk, Peter van Mourik, Anabela S Ramalho, Iris A L Silva, Marvin Statia, Evelien Kruisselbrink, Sylvia W F Suen, Johanna F Dekkers, Frank P Vleggaar, Roderick H J Houwen, Jasper Mullenders, Sylvia F Boj, Robert Vries, Margarida D Amaral, Kris de Boeck, Cornelis K van der Ent, Jeffrey M Beekman

Affiliations

  1. Dept. of Pediatric Pulmonology, Wilhelmina Children's Hospital, University Medical Center Utrecht, Utrecht University, Utrecht, the Netherlands.
  2. Regenerative Medicine Utrecht, University Medical Center Utrecht, Utrecht University, Utrecht, the Netherlands.
  3. Katholieke Universiteit Leuven, Dept. of Development and Regeneration, Leuven, Belgium.
  4. University of Lisboa, Faculty of Sciences, BioISI- Biosystems & Integrative Sciences Institute, Lisboa, Portugal.
  5. Hubrecht Organoid Technology (HUB), Utrecht, the Netherlands.
  6. Prinses Maxima Centrum, Utrecht, the Netherlands.
  7. Gastroenterology and Hepatology, University Medical Center Utrecht, Utrecht, the Netherlands.
  8. Dept. of Pediatric Gastroenterology, Wilhelmina Children's Hospital, University Medical Center Utrecht, Utrecht, the Netherlands.

PMID: 33111074 PMCID: PMC7580120 DOI: 10.1016/j.xpro.2020.100019

Abstract

This protocol describes the isolation, handling, culture of, and experiments with human colon stem cell organoids in the context of cystic fibrosis (CF). In human colon organoids, the function of cystic fibrosis transmembrane conductance regulator (CFTR) protein and its rescue by CFTR modulators can be quantified using the forskolin-induced swelling assay. Implementation procedures and validation experiments are described for six CF human colon organoid lines, and representative CFTR genotypes are tested for basal CFTR function and response to CFTR-modulating drugs. For complete details on the use and execution of this protocol, please refer to Dekkers et al (2016) and Berkers and van Mourik (2019).

© 2020 The Authors.

Conflict of interest statement

J.F.D. is an inventor on a patent related to the organoid technology. M.A.A. reports grants from Gilead Pharmaceuticals, Vertex Pharmceuticals, Proteostasis Therapeutics and personal fees from Vertex

References

  1. Nat Med. 2013 Jul;19(7):939-45 - PubMed
  2. PLoS Biol. 2018 Jul 3;16(7):e2005970 - PubMed
  3. BMC Gastroenterol. 2013 May 20;13:91 - PubMed
  4. Cell Rep. 2019 Feb 12;26(7):1701-1708.e3 - PubMed
  5. Sci Transl Med. 2016 Jun 22;8(344):344ra84 - PubMed
  6. J Vis Exp. 2017 Feb 11;(120): - PubMed

Publication Types