Optimising the decellularization of human elastic cartilage with trypsin for future use in ear reconstruction
Open accessAbstract
Decellularized scaffolds can induce chondrogenic differentiation of stem cells. This study compares different methods to optimise the decellularization of auricular cartilage. The process consisted of an initial 12 hour dry freeze thaw which froze the cartilage specimens in an empty tube at −20 °C. Samples were allowed to thaw at room temperature followed by submersion in phosphate buffer solution in which they were frozen at −20 °C for a 12 hour period. They were then allowed to thaw at room temperature as before. Protocol A subsequently involved subjecting specimens to both deoxyribonuclease and sodium deoxycholate. Protocol B and C were adaptations of this using 0.25% trypsin (7 cycles) and a 0.5 molar solution of ethylenediaminetetraacetic acid (3 hours for each cycle) respectively as additional steps. Trypsin accelerated the decellularization process with a reduction in DNA content from 55.4 ng/μL (native) to 17.3 ng/μL (P-value < 0.05) after 14 days. Protocol B showed a faster reduction in DNA content when compared with protocol A. In comparison to protocol C after 14 days, trypsin also showed greater decellularization with a mean difference of 11.7 ng/μL (P-value < 0.05). Histological analysis with H&E and DAPI confirmed depletion of cells at 14 days with trypsin.
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Mere EDTA treatment is insufficient to achieve suitable acellular ECM scaffolds.
“However, mere EDTA treatment is insufficient to achieve suitable acellular ECM scaffolds [95].”
From Immunogenicity of decellularized extracellular matrix scaffolds: a bottleneck in tissue engineering and regenerative medicine (2023) · cites this paper as [95]
The Results (Histological analysis; page unavailable) report that nuclei remained after 14 days with the EDTA-containing protocol, supporting its insufficiency under those conditions. But Materials and Methods (Decellularization process; page unavailable) show that this protocol also used DNase and other steps, so the study does not establish that EDTA treatment alone is generally insufficient for acellular ECM scaffolds. (LLM, conf=0.86)
Experimental — accuracy not yet measuredTrypsin may impact ECM mechanics.
“Initially used in decellularization for its efficacy in disrupting tissue ultrastructure and enhancing the penetration of other agents, trypsin may nevertheless impact ECM mechanics.”
From Bioactive scaffolds for tissue engineering: A review of decellularized extracellular matrix applications and innovations (2025) · cites this paper as [43]
Shafiq's method for auricular cartilage decellularization integrated SDC, freeze-thaw cycles, DNase, and trypsin, which effectively removed cells and preserved collagen and biomechanical properties.
“Shafiq's method for auricular cartilage decellularization integrated SDC, freeze–thaw cycles, DNase, and trypsin, which effectively removed cells and preserved collagen and biomechanical properties.”
From Bioactive scaffolds for tissue engineering: A review of decellularized extracellular matrix applications and innovations (2025) · cites this paper as [43]
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Research works printed but not linked: 25
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- Authors
- Rahman, S., Griffin, M., Naik, A., Szarko, M., & Butler, P. E. M.
- Affiliations
- University College London (GB)
- The Royal Free Hospital (GB)
- St George's, University of London (GB)
- Corresponding author
- Rahman, Shafiq
- Griffin, Michelle
- Venue
- Scientific Reports
- Published
- 2018-02-15
- Volume
- 8
- Issue
- 1
- Pages
- 3097-3097
- ISSN
- 2045-2322 (ISSN-L)
- PMC
- PMC5814427
- OpenAlex
- W2807223136
- Cited by
- 82
- References
- 28
- Citations by year
- 2026: 4 · 2025: 12 · 2024: 14 · 2023: 12 · 2022: 6 · 2021: 14 · 2020: 9 · 2019: 10 · 2018: 1
- FWCI
- 8.84
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Locations
- Scientific Reports (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
- UCL Discovery (University College London) (submittedVersion)
- Funders
- Action Medical Research
- National Institute for Health and Care Research (ACF-2016-18-022)
- Medical Research Council (MC_PC_15014, MC_EX_MR/M003485/1)
- Topics
- Health Sciences › Medicine › Surgery › Tissue Engineering and Regenerative Medicine
- Physical Sciences › Engineering › Biomedical Engineering › Bone Tissue Engineering Materials
- Physical Sciences › Engineering › Biomedical Engineering › 3D Printing in Biomedical Research
- Keywords
- decellularization, elastic cartilage, auricular cartilage, trypsin, ear reconstruction, decellularized scaffolds, DNase, sodium deoxycholate, EDTA, DNA content, chondrogenic differentiation, freeze-thaw
- MeSH terms
- Cadaver, Cartilage, Cell Differentiation, Ear, Ear, External, Extracellular Matrix, Humans, Stem Cells, Trypsin, Plastic Surgery Procedures, Chondrogenesis, Tissue Engineering, Elastic Cartilage, Tissue Scaffolds, Mesenchymal Stem Cells
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