Journal article
Tunable Hydrogels from Pulmonary Extracellular Matrix for 3D Cell Culture
Journal of Visualized Experiments, Vol.2017(119), e55094
01/17/2017
DOI: 10.3791/55094
PMCID: PMC5352266
PMID: 28117788
Abstract
Here we present a method for establishing multiple component cell culture hydrogels for in vitro lung cell culture. Beginning with healthy en bloc lung tissue from porcine, rat, or mouse, the tissue is perfused and submerged in subsequent chemical detergents to remove the cellular debris. Histological comparison of the tissue before and after processing confirms removal of over 95% of double stranded DNA and alpha galactosidase staining suggests the majority of cellular debris is removed. After decellularization, the tissue is lyophilized and then cryomilled into a powder. The matrix powder is digested for 48 hr in an acidic pepsin digestion solution and then neutralized to form the pregel solution. Gelation of the pregel solution can be induced by incubation at 37 °C and can be used immediately following neutralization or stored at 4 °C for up to two weeks. Coatings can be formed using the pregel solution on a non-treated plate for cell attachment. Cells can be suspended in the pregel prior to self-assembly to achieve a 3D culture, plated on the surface of a formed gel from which the cells can migrate through the scaffold, or plated on the coatings. Alterations to the strategy presented can impact gelation temperature, strength, or protein fragment sizes. Beyond hydrogel formation, the hydrogel stiffness may be increased using genipin.
Details
- Title: Subtitle
- Tunable Hydrogels from Pulmonary Extracellular Matrix for 3D Cell Culture
- Creators
- Patrick A. Link - Virginia Commonwealth UniversityRobert A. Pouliot - Virginia Commonwealth UniversityNabil S. Mikhaiel - Virginia Commonwealth UniversityBethany M. Young - Virginia Commonwealth UniversityRebecca L. Heise - Virginia Commonwealth University
- Resource Type
- Journal article
- Publication Details
- Journal of Visualized Experiments, Vol.2017(119), e55094
- DOI
- 10.3791/55094
- PMID
- 28117788
- PMCID
- PMC5352266
- NLM abbreviation
- J Vis Exp
- ISSN
- 1940-087X
- eISSN
- 1940-087X
- Publisher
- MyJove Corporation
- Language
- English
- Date published
- 01/17/2017
- Academic Unit
- Roy J. Carver Department of Biomedical Engineering; Craniofacial Anomalies Research Center
- Record Identifier
- 9984948142402771
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