Human periodontal ligament cell cultures (hPDL) As described previously (Scanlon et al., 2011), prepare the primary culture of human periodontal ligament cells via the direct cell outgrowth method, by isolating cells from the periodontal ligament tissue around the middle third of healthy human teeth extracted.
{"action": "prepare", "output": "", "reagent": ["human periodontal ligament cells"]}
Maintain cells in approximately 10 mL of MEM-α augmented with 10% FBS, 1% P/S, when they reach approximately 90% confluency using 0.25% trypsin a Thermo Sorvall ST16R refrigerated centrifuge.
{"action": "maintain", "output": "", "reagent": ["MEM-α"], "volume": ["approximately 10 mL"]}
Although variation will be observed from patient to patient, cells usually take approximately three to four days to become confluent after seeding cells at approximately 70%.Use cells .
{"action": "observe", "output": ""}
passaged three to six times for experimentation
{"action": "rinse", "output": "", "volume": ["three to six"]}
Determine enzymatic activity using gelatin zymography as described previously (Cathcart, 2016).Wash cells with PBS 2–3 times.
{"action": "wash", "output": "", "reagent": ["PBS"], "times": ["2–3 times"]}