Universität zu Köln

Fan, Ningbo, Raatz, Lisa, Chon, Seung-Hun ORCID: 0000-0002-8923-6428, Quaas, Alexander, Bruns, Christiane and Zhao, Yue (2022). Subculture and Cryopreservation of Esophageal Adenocarcinoma Organoids: Pros and Cons for Single Cell Digestion. J. Vis. Exp. (185). CAMBRIDGE: JOURNAL OF VISUALIZED EXPERIMENTS. ISSN 1940-087X

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Abstract

The lack of suitable translational research models reflecting primary disease to explore tumorigenesis and therapeutic strategies is a major obstacle in esophageal adenocarcinoma (EAC). Patient-derived organoids (PDOs) have recently emerged as a remarkable preclinical model in a variety of cancers. However, there are still limited protocols available for developing EAC PDOs. Once the PDOs are established, the propagation and cryopreservation are essential for further downstream analyses. Here, two different methods have been standardized for EAC PDOs subculture and cryopreservation, i.e., with and without single cell digestion. Both methods can reliably obtain appropriate cell viability and are applicable for a diverse experimental setup. The current study demonstrated that subculturing EAC PDOs with single cell digestion is suitable for most experiments requiring cell number control, uniform density, and a hollow structure that facilitates size tracking. However, the single cell -based method shows slower growth in culture as well as after re-cultivation from frozen stocks. Besides, subculturing with single cell digestion is characterized by forming hollow structures with a hollow core. In contrast, processing EAC PDOs without single cell digestion is favorable for cryopreservation, expansion, and histological characterization. In this protocol, the advantages and disadvantages of subculturing and cryopreservation of EAC PDOs with and without single cell digestion are described to enable researchers to choose an appropriate method to process and investigate their organoids.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Fan, Ningbo UNSPECIFIED UNSPECIFIED UNSPECIFIED Raatz, Lisa UNSPECIFIED UNSPECIFIED UNSPECIFIED Chon, Seung-Hun UNSPECIFIED orcid.org/0000-0002-8923-6428 UNSPECIFIED Quaas, Alexander UNSPECIFIED UNSPECIFIED UNSPECIFIED Bruns, Christiane UNSPECIFIED UNSPECIFIED UNSPECIFIED Zhao, Yue UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-661689
DOI:	10.3791/63281
Journal or Publication Title:	J. Vis. Exp.
Number:	185
Date:	2022
Publisher:	JOURNAL OF VISUALIZED EXPERIMENTS
Place of Publication:	CAMBRIDGE
ISSN:	1940-087X
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language CARCINOMA; MODELS Multiple languages Multidisciplinary Sciences Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/66168