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Root apical papilla was gently separated from the surface of the root, minced and digested in+ x6 d4 O) n* Z5 m& n/ H
a solution of 3 mg/ml collagenase type I (Worthington Biochemicals Corp., Freehold, NJ) and8 a* Z- e8 g& w6 r9 z
4 mg/ml dispase (Roche Diagnostic/Boehringer Mannheim Corp., Indianapolis, IN) for 30
; n. O. p6 _- k0 h% Jminutes at 37°C. Single cell suspensions of SCAP were obtained by passing through a 70 μm8 Q3 t1 ?" _, y! J
strainer (Falcon, BD Labware, Franklin Lakes, NJ), seeded at 1×104 into 10-cm culture dishes; O2 X- S$ s, ^( C: ^+ n6 w8 A
(Costar, Cambridge, MA), and cultured with α-Modification of Eagle’s Medium (GIBCO/. Q# C; L' m3 P/ n" }
Invitrogen, Carlsbad, CA) supplemented with 15–20% fetal bovine serum (FBS) (Equitech-% T, {/ B# E0 c
Bio Inc., Kerrville, TX), 100 mM L-ascorbic acid 2-phosphate (WAKO, Tokyo, Japan), 2 mM7 a- c6 c) {9 q& e0 q ^, w( Z
L-glutamine (Biosource/Invitrogen), 100 U/ml penicillin and 100 mg/ml streptomycin at 37°# Y" c0 |+ k. u# W
C in 5% CO2. 这是我要养的这个间充质干细胞的培养方法,我想问的是:1:a-MEM和DMEM的区别,当然除了成分上的区别,是不是一个促进细胞的分化?有的人养牙髓干细胞会用GIBCO DMEM/F12(1:1) 11330,有的用hyclone的SH30265.01B。2:100 mM L-ascorbic acid 2-phosphate 这个L-抗坏血酸2-磷酸酯的具体的作用是什么呢?提供vc的营养,只是比vc稳定?可是矿化诱导液里面是加了抗坏血酸的,这不是促进干细胞的分化嘛?3:我按照文献里,查到http://www.wako-chem.co.jp/english/labchem/productinf.htm 这个公司的产品只能查到L-Ascorbic Acid Phosphate Mg Salt.nH2O。sigam公司也只能查到A8960 SIGMA L-Ascorbic acid 2-phosphate sesquimagnesium salt hydrate 。我到底该怎么办呢?这个细胞的培养 我到底该选择什么样的培养基,这个L-抗坏血酸2-磷酸酯 我该怎么买呢? |
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