Culturing Salispheres from Salivary Gland Tissue: A Method to Develop Salispheres from Human Salivary Gland Derived Epithelial Cells
Published: April 30, 2023
Abstract
Source: Beucler, M. J., Miller, W. E. Isolation of Salivary Epithelial Cells from Human Salivary Glands for In Vitro Growth as Salispheres or Monolayers. J. Vis. Exp. (2019)
This video presents the protocol for culturing salispheres, the three-dimensional spheroids derived from salivary glands. The salispheres can be further used to assess the impact of head and neck cancer on saliva production from salivary glands of affected human patients.
Protocol
All procedures involving human participants have been performed in compliance with the institutional, national and international guidelines for human welfare and have been reviewed by the local institutional review board. 1. Tissue Digestion Using a 100 mm tissue culture plate, mechanically mince the tissue into fine pieces ~1–2 mm in size using autoclave-sterilized dissecting scissors and surgical forceps (Figure 1B, C). The…
Divulgaciones
The authors have nothing to disclose.
Materials
| 100 mm culture dishes | Thermo Scientific | 172931 | |
| 15 mL conical tubes | Thermo Scientific | 339651 | |
| 50 mL conical tubes | Thermo Scientific | 339653 | |
| Bronchial Epithelial Cell Growth Media | Lonza | CC-3171 | Add bullet kit as per manufacturer's instructions. Supplement with 20 mL of charcoal stripped serum. |
| Cell strainer 70 µm nylon mesh | Fisher | 22-363-548 | |
| Charcoal stripped fetal bovine serum | Gibco | 12676-029 | |
| Collagenase type III | Worthington | LS004182 | Store at 4 °C. |
| Dispase | Cell Applications | 7923 | Dissolve collagenase to make a 0.15% (w/v) stock. Filter sterilize then store at -20 °C. |
| Dissecting scissors | Fisher | 08-940 | |
| Dulbecco phosphate buffered saline | Corning | 55-031-PC | |
| PathClear Basement membrane extract | Cultrex | 3432-005-01 | Thaw at 4 °C at least 24 hr prior to use. Always handle on ice. |
| Six-well culture dishes | Falcon | 353046 | |
| Surgical forceps | Fisher | 22-079-742 |
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Culturing Salispheres from Salivary Gland Tissue: A Method to Develop Salispheres from Human Salivary Gland Derived Epithelial Cells. J. Vis. Exp. (Pending Publication), e20497, doi: (2023).