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).

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