小鼠骨髓间充质干细胞

viv2005

我做rat的，小鼠的没做过，不知道我的protocol有参考价值没，感觉MSC是最好培养的了，比什么osteoblast和osteoclast好养多了
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MSC  Isolation from Rat Femur and Tibia
8 \+ S9 i- R5 y2 l0 E* E4 X( _5 w1.        Euthanize the rats with CO2 for 5 mins.
1 P( o( {+ g( s; I2.        Isolate femur and tibia by disconnecting them at the joints.
/ i% l; {- {* v3.        Clean the bones of soft tissues.
& o5 ]$ m; e- p$ b4 V4.        Wash both femur and tibia in PBS containing 1% penicillin/streptomycin twice.
" b8 o7 J9 X, B. C$ o" y# v5.        Under the cell culture hood, clip-off the epiphyseal plates to expose the bone marrow., o( [5 O2 N7 z7 n% _5 \8 ?5 Y
6.        Using a syringe, flush the bone marrow with 10 ml of media into a dish.
* E0 K$ w, E, [3 Y2 r+ B. w7.        Break up the clumps by aspirating the cell solution multiple times.. |) {9 h3 x+ F4 v& O
8.        Using a 70μm cell strainer, filter the cell solution into 15ml tube. This will remove any unwanted debris and bone fragments.
$ Q; X# A- O- A5 A$ G9.        Centrifuge the filtered cell solution at 1000rpm for 10 min.& a' b7 y6 M4 Z
10.        Resuspend the pellet and pipet this solution into a 100mm dish.
* e. {( t$ w7 a# o* P% ~11.        Incubate for the next 48h.' M$ S# G$ \; D# Q1 {
12.        Change media to remove non-adhering cells.
& a5 u' e0 X2 L, X+ G4 L: Q# c& k13.        Change media every 2-3 days, even the media doesn’t change the color.
) x4 T( e$ v- }. R! d14.        Usually the cell will be confluence at 7-10 days, and then are ready for passaging.7 l( Y# X/ S+ j% W1 b7 }5 W