Isolation of Gingival Immune Cell Network from Murine Teeth Blocks
Published: August 31, 2023
Abstract
Source: Dutzan, N. et al., Isolation, Characterization and Functional Examination of the Gingival Immune Cell Network. J. Vis. Exp. (2016)
This video demonstrates the isolation procedure of the gingival immune cell network from a mouse model. The enzymatic digestion releases the gingival tissue from the attached teeth and bone, which is further mechanically digested, releasing immune cells from the gingiva. The isolated cells are ready for immunological experiments.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Prepare in Advance Prepare Complete media: RPMI supplemented with 2 mM L-glutamine, 100 units/ml penicillin, 100 ug/ml streptomycin, and 10% FBS. Prepare DNase media: 50 ml of RPMI media supplemented with 7.5 µg DNase (Make fresh, keep on ice at all times). Prepare Collagenase-DNase media: 5 ml …
Representative Results
Figure 1: Isolation of murine gingiva. Illustration demonstrating the borders of dissection in the maxilla and the isolated gingival segment to be used for gingival tissue dissection. (B) Segment of the murine maxilla (molar area) with an outline for block dissection, and isolated block. Hematoxylin and Eosin stain (H&E) of a maxillary molar segment with gingival tissues outlined is shown in low (C) and higher magnifica…
Divulgaciones
The authors have nothing to disclose.
Materials
| Fine Scissors | Fine science tools | 14058-11 | |
| Scalpel Handle #3 | Fine science tools | 10003-12 | |
| Scalpel Blades #10 | Fine science tools | 10010-00 | Sterile |
| Splinter Forceps | Integra Miltex | 6-304 | |
| Needles with regular bevel | BD Medical | 305109 | 27G, 12.7 mm length |
| Monoject syringes | Covidien | 8881513934 | Luer-lock tip, 3mL |
| PBS, pH 7.4 | Life Technologies | 10010-049 | Without Calcium and Magnesium |
| RPMI 1640 | Lonza | 12-167F | Without L-glutamine |
| DNase I from bovine pancreas | Sigma-Aldrich | DN25-1G | |
| Collagenase type IV | Gibco (by Life technologies) | 17104-019 | |
| Fetal Bovine Serum | Gemini Bio-products | 100-106 | |
| Gentamicin 50 mg/ml | Quality biological | 120-098-661EA | |
| Pen Strep | Gibco (by Life technologies) | 15140-122 | |
| L-Glutamine | Gibco (by Life technologies) | 25030-081 | |
| 0.5M EDTA pH 8.0 | Quality biological | 351-027-721EA | |
| 50 mL tubes | Corning | 352070 | Polypropylene, sterile |
| 70 μM Cell Strainers | Corning | 352350 | |
| Petri dishes | Corning | 351029 | Sterile |
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Citar este artículo
Isolation of Gingival Immune Cell Network from Murine Teeth Blocks. J. Vis. Exp. (Pending Publication), e21566, doi: (2023).