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发表于 2012-1-13 13:55 |只看该作者 |倒序浏览 |打印

本帖最后由细胞海洋于 2012-1-13 14:38 编辑 - F1 `$ {+ G9 e. [
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Springer出版社的新书，很多图，对做人多能干细胞的童鞋可能有用。0 [! j; a) S  K0 h+ E, n
ISBN 978-1-61779-547-3 e-ISBN 978-1-61779-548-0$ [/ V- R- f& f5 |& W
DOI 10.1007/978-1-61779-548-0
Springer New York Dordrecht Heidelberg London
Library of Congress Control Number: 20119416080 f9 D1 W+ z* u/ b0 ]
© Springer Science+Business Media, LLC 20129 j9 R/ @0 d" y" v/ z/ q
[hide][/hide]
Contents：
1Methods for the Derivation of Human Embryonic
Stem Cell Lines .......................................................................................... 1
1.1 Introduction ........................................................................................ 1
1.2 Materials for ESC Line Derivation .................................................... 9
1.3 Methods for hESC Isolation .............................................................. 9
1.3.1 hESC Isolation by Immunosurgery........................................ 12
1.3.2 Mechanical Removal of Trophectoderm ................................ 12
1.3.3 Whole Embryo Approach for ESC Line Derivation .............. 131 x* w; M' }! U+ \0 Y3 ?: O8 P
References ................................................................................................... 13" A( M7 `& O( F0 B2 A3 S, W
2 Morphology of Human Embryonic and Induced Pluripotent5 B% N  Q6 a4 x* j. g; i$ u
Stem Cell Colonies Cultured with Feeders ............................................. 15
2.1 Introduction ........................................................................................ 15
2.2 Materials ............................................................................................ 167 i7 [% a4 G; k; g- U* W
2.2.1 For Mouse Embryonic Fibroblasts (MEFs)4 s* P+ F( T2 ?5 Z; H1 j" ^. u: ~7 s
and Foreskin Fibroblasts (HFFs) ........................................... 16
2.2.2 For hPSC Maintenance .......................................................... 178 o$ y1 Q! K4 g. `9 F
2.3 Methods ............................................................................................. 18) u& D% B5 _1 F: p; w9 u
2.3.1 Feeder Culture Methods ........................................................ 18% ]4 i/ a+ H' s7 F: X
2.3.2 hPSC Culture ......................................................................... 22; w: o- A' M- p# b- t! E, H4 E
References ................................................................................................... 385 r1 N+ |+ Y, T7 ]3 y1 W/ G- ^! M
3 Morphology of Human Embryonic Stem Cells and Induced1 j& h6 z) i5 J0 l) {, t
Pluripotent Stem Cells Cultured in Feeder
Layer-Free Conditions .............................................................................. 41
3.1 Introduction ........................................................................................ 41
3.2 Materials for Feeder Layer-Free Culture of hPSCs ........................... 43
3.2.1 Matrix Preparation ................................................................. 43
3.2.2 Culture Medium ..................................................................... 443 b  x) |6 N; Z
3.3 Methods for hPSC Feeder Layer-Free Culture .................................. 44
3.3.1 Preparation of Matrix-Covered Plates ................................... 44
3.3.2 Splitting, Freezing, and Thawing hPSCs ............................... 45' r. O. V/ ^8 M, m1 N) n
3.3.3 Adaptation of PSCs to Feeder-Free Culture .......................... 45
3.3.4 Routine Culture of hPSCs ...................................................... 46
References ................................................................................................... 54; X7 U6 i- u6 r1 J6 t
4 Morphology of Undifferentiated Human Embryonic/ ]7 D# w& W$ o9 z
and Induced Stem Cells Grown in Suspension
and in Dynamic Cultures .......................................................................... 57
4.1 Introduction ........................................................................................ 57
4.2 Materials for Suspension Culture of hPSCs ...................................... 58/ Q. q( r/ M: N8 r# w3 d
4.2.1 Culture Medium ..................................................................... 58
4.2.2 Splitting Medium ................................................................... 59, K& N6 c1 n( w) N
4.2.3 Freezing Medium ................................................................... 59
4.3 Methods for Suspension Culture of hPSCs ....................................... 60
4.3.1 Creating a hPSC Suspension Culture .................................... 60
4.3.2 Splitting hPSCs in Suspension ............................................... 60$ ?6 A9 w; z8 [6 }' B$ {* C4 S
4.3.3 Freezing hPSCs in Suspension .............................................. 62
4.3.4 Thawing hPSCs in Suspension .............................................. 64
4.3.5 Culturing hPSCs in a Dynamic System ................................. 64
4.3.6 Routine Culture of hPSCs in Suspension .............................. 65( W) A1 L! A6 A) c
References ................................................................................................... 719 y0 }& f  l( l* x/ _. C
5 Differentiation of Pluripotent Stem Cells In Vitro:
Embryoid Bodies ....................................................................................... 735 Y7 A* _- L; F9 }) r
5.1 Introduction ........................................................................................ 73. g; a; U( }' k) K: \% [  A, j" R& I, M
5.2 Materials for EB Formation ............................................................... 75
5.2.1 Culture Medium Supplemented with Serum ......................... 75" g4 z' R5 h% H  q0 Q3 l) N" @
5.2.2 Splitting Medium Based on Collagenase ............................... 766 P/ `% y4 L$ v) v2 z
5.3 Methods for EB Formation and Culture ............................................ 76, |  c" e  c0 w0 W! g* [- Z! G- v
5.3.1 EB Formation ......................................................................... 76
5.3.2 Routine Culture of EBs .......................................................... 76
5.3.3 Culturing EBs in Spinner Flasks ............................................ 784 q: V; S/ z" A! d. Z) D
References ................................................................................................... 88) ~& P( p+ U& z$ G6 g) Z
6 Differentiation of Pluripotent Stem Cells In Vivo:
Teratoma Formation ................................................................................. 91$ C) m4 z; L0 z' l7 R5 u, r! l0 n
6.1 Introduction ........................................................................................ 91
6.2 Materials for Teratoma Formation ..................................................... 93
6.2.1 Culture Medium ..................................................................... 93
6.2.2 Syringe for Injecting Cells ..................................................... 93
6.3 Formation of Teratomas ..................................................................... 93; [% K- ?! K$ j& N
6.3.1 Protocol for Teratoma Formation .......................................... 938 X) \0 v' P+ P7 p% H" h8 x
6.3.2 Routine Treatment of Mice and Teratoma ............................. 93
References ................................................................................................... 103! B, j) \) i! Z
7 Immunostaining ........................................................................................ 105
7.1 Introduction ........................................................................................ 105
7.2 Materials and Solutions for Immunostaining .................................... 111
7.2.1 Materials and Solutions for Immunohistochemistry, i: p6 r6 a0 h6 z7 ?$ `
of Paraffi n-Embedded Tissues ............................................... 111
7.2.2 Materials and Solutions for Immunofl uorescence ................. 111
7.3 Immunostaining Procedures .............................................................. 112! U( x3 Q  t$ g2 E# Z. z# f
7.3.1 Immunohistochemistry of Paraffi n-Embedded Tissues ......... 112" W& C5 }; s  s
7.3.2 Immunofl uorescence of Cultured Cells ................................. 113
References ................................................................................................... 113
8 Karyotype and Fluorescent In Situ Hybridization1 h9 p7 r1 F% r# |
Analysis of Human Embryonic Stem Cell and Induced
Pluripotent Stem Cell Lines ..................................................................... 115
8.1 Introduction ........................................................................................ 115* N, C" n* Y5 p% F4 h$ K  s. g
8.1.1 Karyotype Analysis ............................................................... 115( V5 U# x* s. c& u. v" t$ _4 B% e& o
8.1.2 FISH Analysis ........................................................................ 121
8.2 Materials for Harvesting Cells for Karyotyping& P3 r& V+ I8 ]1 s/ m( W2 x" ]2 l
and FISH Analysis ............................................................................. 1241 n7 `, w. h) B; n: j- T* w1 f
8.2.1 Reagents ................................................................................. 124' y2 L( h- z) l/ C/ ^0 p4 B
8.2.2 Solutions ................................................................................ 124% |% A" s& u  q: i+ n7 D3 u- u
8.3 Procedure of Harvesting Cells for Karyotyping
and FISH Analysis ............................................................................. 1244 ^: s+ p- I. q3 F/ _3 J: N/ d7 ^
References ................................................................................................... 126
9 Method for the Derivation of Induced Pluripotent
Stem Cells from Human Hair Follicle Keratinocytes ............................ 1276 j4 v5 o" H, m$ D7 g+ a
9.1 Introduction ........................................................................................ 1276 f; z% M0 P# E, |  D, \% @" P$ a
9.2 Materials ............................................................................................ 129
9.2.1 NIH-3T3/293T Cells .............................................................. 129( z) a9 N( M9 s$ a# K- W
9.2.2 Keratinocyte Derivation from Plucked Hair Follicles ........... 129
9.3 Methods ............................................................................................. 130( @) X7 _4 W& @) W/ p  H
9.3.1 NIH-3T3 and 293T Culture Methods .................................... 130( r. M5 j7 r& ^
9.3.2 Keratinocyte Culture Methods ............................................... 132
9.3.3 Preparation of the STEMCCA Virus for Infection ................ 133' m$ {, z$ ]! a% A- H! B
9.3.4 Derivation of iPSCs from Hair Keratinocytes ....................... 1344 }1 U' F0 F+ ]/ k
References ................................................................................................... 137

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