Isolation of Circumvallate Papillae (CVP) Epithelium from Mouse Model: An Enzymatic Procedure to Separate CVP Epithelium from Mouse Tongue
Trascrizione
The dorsal surface of the mouse tongue is covered with three main kinds of raised protrusions called papillae, namely fungiform, circumvallate, and filiform papillae. The circumvallate papillae, or CVP, comprising epithelial trenches that house taste buds, is supported by a layer of connective tissue – the lamina propria.
To isolate the CVP epithelium, begin with a euthanized mouse in the supine position. Next, incise the cheeks and dissect the thick band of tissue – the lingual frenulum – to separate the tongue from the floor of the oral cavity. Transfer the excised tongue into a dish containing chilled buffer.
Next, cut out the anterior part of the tongue. Carefully inject a digestion cocktail containing dispase and collagenase enzymes into the subepithelial space of the CVP region, between the epithelium and underlying tissues. Retract the needle while injecting the suspension into the region.
Incubate the tongue for the appropriate duration. Dispases and collagenases digest the extracellular matrix proteins within the connective tissue, facilitating the separation of CVP epithelium from the tissue. Following incubation, make incisions in the epithelium in the CVP region.
Gently separate the loosened epithelium from the underlying tissue. Finally, transfer the separated CVP epithelium into a protein-coated tube to prevent tissue from sticking to the tube wall. The CVP epithelium can be stored for further downstream processing.
