MSC成脂成骨诱导图片

Learner

回复 皮搋子 的帖子' j0 V. m* z/ T
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1uM=1000nM

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回复 stream 的帖子
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$ s# u1 B# g/ `0 Q- M" y0 U9 i  Q干细胞大部分都分化了，有些未分化的也老化了，我用的细胞不是经过流式细胞仪筛选的，有些本身就不是干细胞。

lkzjyx

请问楼主是做的哪种动物的MSC呢？用的什么方法分离和培养的，谢谢   " d% c3 E" k8 u
因为我也在做这个方面的研究  我做的是犬的BMSC 想跟你交流下

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回复 luoziwei 的帖子. b: z' z5 N9 T' m4 B, U
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很抱歉回复不及时，这几天出门了。
; u2 n+ S& c) N, C" j! L下面是我的染色方法：. }, R; h5 M. c! b
固定前细胞吸去培养基用PBS洗一到二次。2 c2 x; e1 p! J1 j8 {2 k5 A& ?
Alizarin Red S staining:( t$ \# a! w1 \1 M! R
1. 4%PFA fixation at RT for 30min.& V* f# w8 l' C2 {! D# h
2.Then wash wish PBS twice.$ r+ W2 V+ B; R/ H" T: P  o8 h
3. Add 1ml of 2% alizarin red water solution (pH4.1-4.3 use 1% NH4OH adjust pH) stain RT for 20min with gentle shaking on a rocker.
# K/ C* }' [2 r1 q& B4.Then wash with PBS for 4 times with gentle shaking. The washing time could be up to 1 hour.8 \% v/ @. p4 a3 ~9 L9 v2 m; q
5.Leave some PBS in the well and look under microscope.

皮搋子

多少毫升中加1uM？

Learner

回复 皮搋子 的帖子6 U& k9 ~( n5 L! w% G
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1uM=1umol/L 这要取决于你在诱导中的终体积。
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Learner

回复 luoziwei 的帖子
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关于油红O染色步骤：
; O# I: M7 \4 O  _5 a- v1。 Cell fixation with 4%PFA RT for 30min.
" a% r2 `& m# ?7 m" k- C0 [7 N% U2.   Remove PFA and wash cell with PBS once./ J6 O( I6 g$ h4 t  L6 ^" {; v' |
3. Then wash cell with 60% isopropanol for 30 min on a shaker.
; O& p. r8 ]& K% Y0 s4. Remove 60% isopropanol and add working oil red O solution stain for 30 min on a shaker.
6 ]. u# t- Y0 D7 Y9 [: O5. After 30min remove oil red O stain immediately and wash with PBS for 4 times.
$ f: h" K: f8 E5 e) [+ Z6. Leave some PBS in the well and take picture.
# G" i9 p) D8 q; `6 {The working oil red solution should be made fresh from the stock each time.
+ F; g$ h5 [$ Q2 r. O9 X7 v8 oStock solution (175mg oil red O/50ml 100% isopropanol) could store for up to one year.
; H4 W! M, s9 M9 DWorking solution: dilute the oil red O stock solution with PBS(6ml oil red O stock+4ml PBS) filter with filter paper or 0.45um filter unit.

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回复 lkzjyx 的帖子
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% j& i+ i8 {: b9 V! D4 W) khuman bone marrow MSC

hxf_86

这个图片很好啊~学习了~

luoziwei

回复 Learner 的帖子/ a0 Y# `: p, m
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有两个小问题想请教一下，你是直接在孔板里诱导染色的？还是把细胞种在玻片上然后取出玻片进行染色拍照的？另外一个问题是，关于油红0染色，用60%的异丙醇洗涤是为什么呢？