Induction induction

Experimental operation process:
osteogeneous induced culture fluid equipped with induction:
1. Prepare a A-MEM culture solution containing 10% FBS;
2 In the amount of EM culture Add 5OUM Decreasic acid 10mm micro-glycerol 00m dexamethasone;
3, prepare 6-bonus culture plate, inoculated P4 cells in specification of 5 * 103 square gravity rice
EM In the culture solution;
4, after the cells are long to substantially fuse,
replaces the prepared osteogeneous induced culture solution;
5, once every three days, the cells are concentrated for 7 days for alkali Sexual research tuning and tuning;
6, twenty-eight days later, mineralized nunasin red dyeing.
Preface:
1. Removal of the cells Current use of culture medium, use PBS to clean twice;
2 Use 10% formaldehyde room temperature for 15 minutes, after completion, use heavy steamed water twice
3 Follow 1 ml of the wind to add 40mm Cao Soil-dyed wave, incubate at room temperature and slightly sway;
4, remove the dye without completely binding, wash the water ticket washed and oscillated 5 minutes Repeat 4 Second;
5, the tilt is placed 2 min to absorb excess returning warm water: inverted the micro-town observation. u0026 rsquo;
Yipin-induced culture fluid equipped with induction operation:
1, configure FBS10% over Hg DMEM culture solution;
, in restricted C DMIE. Dieni Dieni Semetzon 1U / M insulin, 20 o'clock, Thill Sixin and 0 5mm buxx,
3, with a 6-hole culture plate, inoculated P4 cells in accordance with 2x104 square centimeter specification in
pump hg- DMEM culture fluid
small to be cellulated to be substantially fused, replace it. The above-mentioned preparative completed ylen-induced culture wave culture is 2 days,
maintains culture liquid culture
moisture with only 10 ug / m1 insulin, so alternate circulation to 1 day, using red oil 0 dyeing .
Red Oil 0 Dyeing Method:
1. Remove the current culture medium used by cells, use PBS to clean twice;
2, 27 .5% formaldehyde room temperature for 20 minutes;
3, heavy steaming water cleaning 3 This, dry in the air;
4, add 0.5% red oil at room temperature - - hour;
5, formulated 70% ethanol solution 3 times;
6. Invert the microscope to observe the photo record.