Determine cell count using a Casy cell counter . #####%%%%% output "cell count" are used in line8
{"action": "determine", "output": "cell count", "device": ["Casy cell counter"]}
Mix 25 µl Opti-MEM medium with 0.5 µg HyPerER plasmid #####%%%%% output "Fugene® HD solution" are used in line6
{"action": "mix", "output": "Fugene\u00ae HD solution", "volume": ["25 µl"], "mass": ["0.5 \u00b5g HyPerER"]}
1.5 µl Fugene® HD solution, incubate for 15 min at room temperature, .
{"action": "incubate", "output": "", "reagent": ["Fugene\u00ae HD solution"], "temperature": ["room temperature"], "time": ["15 min"]}
add to cells #####%%%%% output "cells" are used in line12
{"action": "add", "output": "cells", "container": ["cells"], "reagent": ["cell count"]}
Add 2.6 ml growth medium 6 h after transfection, .
{"action": "add", "output": "", "volume": ["2.6 ml"], "reagent": ["growth medium"]}
incubate cells at 37°C for 24-48 h. 8
{"action": "incubate", "output": "", "reagent": ["cells"], "temperature": ["37\u00b0C"], "time": ["24-48 h"]}
Perform imaging on an Olympus Fluoview 3000 microscope with temperature control. #####%%%%% output "culture medium" are used in line16
{"action": "perform", "output": "culture medium", "device": ["Olympus Fluoview 3000 microscope"]}
Remove culture medium, , . #####%%%%% output "pre-warmed HEPES Imaging Buffer" are used in line18
{"action": "remove", "output": "pre-warmed HEPES Imaging Buffer", "reagent": ["culture medium"]}
wash dish twice with 1 ml pre-warmed HEPES Imaging Buffer
{"action": "rinse", "output": "", "volume": ["1 ml"], "reagent": ["pre-warmed HEPES Imaging Buffer"]}
add 1 ml HEPES Imaging Buffer, place dish on microscope stage #####%%%%% output "stimulating substances (e.g., thapsigargin)" are used in line22
{"action": "add", "output": "stimulating substances (e.g., thapsigargin)", "volume": ["1 ml HEPES Imaging Buffer"], "device": ["microscope stage"], "reagent": [""]}
Measure baseline oxidative status every 20 s for at least 5 min, add stimulating substances (e.g., thapsigargin) . #####%%%%% output "baseline oxidative status" are not used
{"action": "add", "output": "baseline oxidative status", "reagent": ["stimulating substances (e.g., thapsigargin)"]}
Conclude measurement with 100 µM H2O2 as a positive control. #####%%%%% output "measurement" are not used
{"action": "proceed", "output": "measurement", "reagent": ["100 \u00b5M H2O2"]}