Transfect HCC cells with ISX fusion GFP expression plasmid select stable clones. #####%%%%% output "stable clones" are used in line14
{"action": "transfect", "output": "stable clones", "reagent": ["ISX fusion GFP expression"]}
Grow clones in MEM culture medium with 10% FBS .
{"action": "generate", "output": "", "reagent": ["clones"]}
Prepare 0.6% , .
{"action": "prepare", "output": "", "concentration": ["0.6% agar"]}
keep warm at 65°C
{"action": "keep", "output": "", "temperature": ["65\u00b0C"]}
If the agar solidifies, repeat the preparation steps. #####%%%%% output "the preparation steps" are not used
{"action": "repeat", "output": "the preparation steps", "time": [""]}
Harvest cells, dilute to 1 x 10^4 cells/ml in 2x MEM with 20% FBS.
{"action": "dilute", "output": "", "concentration": ["1 x 10^4 cells/ml", "20% FBS"]}
Mix 1 ml diluted cells with 1 ml 0.6% agar, solidify at 37°C for 30 min.
{"action": "mix", "output": "", "concentration": ["0.6% agar"], "reagent": ["stable clones"]}
Add 2 ml 10% FBS MEM culture medium to each well. #####%%%%% output "the medium addition" are used in line18
{"action": "add", "output": "the medium addition", "volume": ["2 ml"], "concentration": ["10% FBS MEM culture medium"]}
Repeat the medium addition every 3 days.
{"action": "repeat", "output": "", "reagent": ["the medium addition"]}
If the staining is not clear, repeat the staining process. #####%%%%% output "the staining process" are not used
{"action": "repeat", "output": "the staining process", "time": [""]}