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( `3 x0 a) d; J. e; Y3 tCurrent Protocols in Stem Cell Biology
# p; w5 N; C* x! S2 } z, vLast Update: July 07, 20099 B( x7 q+ n# p) ]& ~
Print ISSN: 1941-7322
2 F- r# B, N* a4 _Online ISSN: 1938-89695 v. B0 F1 c( Q/ y4 A
Table of Contents! e5 a% v; g q- i ?1 K
Foreword
' \' G6 t, O( N+ z; j. TPreface , }; R* {2 R# b
Chapter 1 Embryonic and Extraembryonic Stem Cells
' S ~5 `" I- X' e8 f9 _ Section A Isolation of Embryonic Stem Cells ; A* B0 Q' [$ h8 p, g
Isolation of Embryonic Stem Cells % l& K* R- I8 N: n! N$ T! J/ q
UNIT 1A.1 Derivation and Characterization of Nonhuman Primate Embryonic Stem Cells - v* B' }) {. \$ w: n0 ]# K
UNIT 1A.2 Derivation of hESC from Intact Blastocysts
7 v$ d6 K- x* ?; u% u- y4 ^' {6 tUNIT 1A.3 Reprogramming Primordial Germ Cells (PGC) to Embryonic Germ (EG) Cells Y- A6 i/ _6 U1 E) G
UNIT 1A.4 Derivation and Propagation of hESC Under a Therapeutic Environment ( ~. v6 c! M9 ^. x4 ]- o5 x
Section B Characterization of Embryonic Stem Cells 0 n' @ Q$ O( f" w
UNIT 1B.1 Proteomic Analysis of Pluripotent Stem Cells ; g4 V+ _. m$ Y6 @' o4 J# w/ U" `
UNIT 1B.2 Gene Expression Analysis of RNA Purified from Embryonic Stem Cells and Embryoid Body–Derived Cells Using a High‐Throughput Microarray Platform
: S; R4 @6 h1 Z5 I' P9 uUNIT 1B.3 Phenotypic Analysis of Human Embryonic Stem Cells - n, R" ?8 n0 D: \& r/ s
UNIT 1B.4 Isolation of Human Embryonic Stem Cell–Derived Teratomas for the Assessment of Pluripotency % _, y7 [; u/ i5 J0 h2 k, B
UNIT 1B.5 Tandem Affinity Purification of Protein Complexes in Mouse Embryonic Stem Cells Using In Vivo Biotinylation * R; l8 E8 F6 r. A! ?9 Z0 J3 w# F
Section C Culture and Maintenance of Undifferentiated Embryonic Stem Cells 2 P0 `; S) S' _6 E
Culture and Maintenance of Undifferentiated Embryonic Stem Cells
, K2 g( P, o0 t/ tUNIT 1C.1 Expansion of Human Embryonic Stem Cells In Vitro
0 N3 O$ [" ?2 }0 DUNIT 1C.2 Defined, Feeder‐Independent Medium for Human Embryonic Stem Cell Culture ; A/ d$ H, r# a, p, f. f" ]
UNIT 1C.3 Isolation and Propagation of Mouse Embryonic Fibroblasts and Preparation of Mouse Embryonic Feeder Layer Cells
" J/ e1 ?. q( y( jUNIT 1C.4 Culture of Mouse Embryonic Stem Cells . M/ Z4 J8 L: }, k! u
UNIT 1C.5 Preparation of Autogenic Human Feeder Cells for Growth of Human Embryonic Stem Cells
4 T. W$ V* K7 u8 a F+ W9 [" hUNIT 1C.6 Isolation of Human Placental Fibroblasts . o3 D" V# p$ X1 w @: P9 v
UNIT 1C.7 Derivation of Human Skin Fibroblast Lines for Feeder Cells of Human Embryonic Stem Cells
( {9 f" n4 G! k, s* g% U4 z9 nUNIT 1C.8 Cryopreservation of Dissociated Human Embryonic Stem Cells in the Presence of ROCK Inhibitor + @. L8 V/ v5 `# {+ F$ f
Section D Germ Layer Induction/Differentiation of Embryonic Stem Cells
9 l3 l0 Z) Z' I5 b& iUNIT 1D.1 Germ Layer Induction in ESC—Following the Vertebrate Roadmap ! U5 B1 i! L0 ~9 t G7 ?
UNIT 1D.2 Formation and Hematopoietic Differentiation of Human Embryoid Bodies by Suspension and Hanging Drop Cultures 6 E6 ]& n5 l' D& H0 i9 [
UNIT 1D.3 Directed Differentiation of Human Embryonic Stem Cells as Spin Embryoid Bodies and a Description of the Hematopoietic Blast Colony Forming Assay " d- M7 }2 ^2 o: Q- C
UNIT 1D.4 Differentiation of Human Embryonic Stem Cells in Adherent and in Chemically Defined Culture Conditions ' d0 O5 Z1 A! i
UNIT 1D.5 Isolation and Differentiation of Xenopus Animal Cap Cells 7 _, y% }+ O. h- L
Section E Extraembryonic Lineages
3 o4 M5 g& n+ R0 {. Z1 bIsolation of Stem Cells from Extraembryonic Tissues , D# O! F& C% E4 l4 G ]
UNIT 1E.1 Isolation of Human Placenta‐Derived Multipotent Cells and In Vitro Differentiation into Hepatocyte‐Like Cells
6 q0 y$ ~# F9 CUNIT 1E.2 Isolation of Mesenchymal Stem Cells from Amniotic Fluid and Placenta
: T y) [' {5 ]) @" Q0 YUNIT 1E.3 Isolation of Amniotic Epithelial Stem Cells + S6 C) \! D) q }* N& y
UNIT 1E.4 Isolation and Manipulation of Mouse Trophoblast Stem Cells & |9 h' a Y+ i) ~& k6 O
Section F Mesodermal Lineages
! i- ]& Z1 w2 tUNIT 1F.1 Differentiation of Embryonic Stem Cells into Cartilage Cells
, ^0 Z; |: h2 `- W4 t! A- I" I% rUNIT 1F.2 Differentiation of Human Embryonic Stem Cells to Cardiomyocytes by Coculture with Endoderm in Serum‐Free Medium
/ N8 Y4 O) z# t9 J4 C' k& T5 \& {UNIT 1F.3 Isolation of Hematopoietic Stem Cells from Mouse Embryonic Stem Cells
2 V5 v; B) _7 q4 b! M% bUNIT 1F.4 Differentiation of Mouse Embryonic Stem Cells into Blood - Y. X& ?1 d4 g- W4 N0 P0 a
UNIT 1F.5 Endothelial Differentiation of Embryonic Stem Cells # R& t% F5 v; E7 O
UNIT 1F.6 Hematopoietic Differentiation of Human Embryonic Stem Cells by Cocultivation with Stromal Layers / j1 a% \, O$ t# D0 D
UNIT 1F.7 TLX1 (HOX11) Immortalization of Embryonic Stem Cell–Derived and Primary Murine Hematopoietic Progenitors
! Y% O3 f$ e/ i9 [. D$ V) D. uUNIT 1F.8 Differentiation of Multipotent Mesenchymal Precursors and Skeletal Myoblasts from Human Embryonic Stem Cells 8 K) O% ^7 l7 \" W. F4 W. Y1 k6 O
Section G Endodermal Lineages : q' R6 z; ?# m/ m/ p( d O4 a
UNIT 1G.1 The Differentiation of Distal Lung Epithelium from Embryonic Stem Cells
* n( f' G3 M: }4 v$ p7 rUNIT 1G.2 Pancreas Differentiation of Mouse ES Cells
& k! F. w% d& z0 i! sUNIT 1G.3 Differentiation of Embryonic Stem Cells into Anterior Definitive Endoderm 0 b4 m& k: U5 Y6 c
Section H Ectodermal Lineages 8 i6 m! H( ?3 a$ T! y+ j
UNIT 1H.1 Differentiation of Mouse Embryonic Stem Cells to Spinal Motor Neurons
) S/ E3 ^0 R' D, E* L Chapter 2 Somatic Stem Cells & a& |" F# s' i9 Z$ i: n5 |
Section A Hematopoietic Stem Cells + V! U# Z3 F; f) j
Hematopoietic Stem Cells
+ U, P) ^9 ^ y2 w, ^UNIT 2A.1 Isolation of Mononuclear Cells from Human Cord Blood by Ficoll‐Paque Density Gradient
w* g. [3 G9 L/ w5 fUNIT 2A.2 Isolation of Hematopoietic Stem Cells from Human Cord Blood $ m" [0 \6 j6 U: n& o! J
UNIT 2A.3 Isolation of Mesenchymal Stem Cells from Human Cord Blood
! h, G6 e& O: y# ~# t1 A( _# yUNIT 2A.4 Isolation and Assessment of Long‐Term Reconstituting Hematopoietic Stem Cells from Adult Mouse Bone Marrow }1 g6 X3 N- \
UNIT 2A.5 Analysis of the Hematopoietic Stem Cell Niche 8 z# C3 e$ o; N! U7 g! o* P0 I
UNIT 2A.6 Analysis and Manipulation of Hematopoietic Progenitor and Stem Cells from Murine Embryonic Tissues
2 j2 n, q5 G+ Z! v; ]8 u4 X3 {$ h7 ]UNIT 2A.7 High Level In Vitro Expansion of Murine Hematopoietic Stem Cells
" J0 N# U. ?; I+ ZUNIT 2A.8 Isolation and Visualization of Mouse Placental Hematopoietic Stem Cells - O2 y8 M3 c# i" b
Section B Non‐Hematopoietic Bone Marrow‐Derived Stem Cells % F+ O j/ {! L. B3 j+ C
UNIT 2B.1 Isolation and Characterization of Mesoangioblasts from Mouse, Dog, and Human Tissues , A# u- N1 x* v+ f
UNIT 2B.2 Purification and Culture of Human Blood Vessel–Associated Progenitor Cells
2 v0 H, Y, H8 h6 ~4 i* a) O; pUNIT 2B.3 Isolation, Culture, and Differentiation Potential of Mouse Marrow Stromal Cells
. r2 h- N0 H' h3 x% l$ H Section C Cardiovascular Stem Cells
6 A z8 G3 W6 O9 nUNIT 2C.1 Isolation and Characterization of Endothelial Progenitor Cells from Human Blood
0 [6 T( A e" QUNIT 2C.2 Derivation of Epicardium‐Derived Progenitor Cells (EPDCs) from Adult Epicardium
4 l( v8 J- g. y8 P! k [ Section D Neural Stem Cells 8 s' u& ?- X5 ^0 _% R
UNIT 2D.1 Generation of Cultured Oligodendrocyte Progenitor Cells from Rat Neonatal Brains 4 N7 |, b/ q( K
UNIT 2D.2 Isolating, Expanding, and Infecting Human and Rodent Fetal Neural Progenitor Cells 9 F& k! t3 U$ L
UNIT 2D.3 Long‐Term Multilayer Adherent Network (MAN) Expansion, Maintenance, and Characterization, Chemical and Genetic Manipulation, and Transplantation of Human Fetal Forebrain Neural Stem Cells + i3 [2 l0 U+ b- M
Chapter 3 Cancer Stem Cells
/ i5 Q4 d+ N# V& g! ?0 oUNIT 3.1 Colon Cancer Stem Cells 5 `9 |$ O) A( D, E# Y r, _
UNIT 3.2 In Vivo Evaluation of Leukemic Stem Cells through the Xenotransplantation Model , N$ ~8 [1 Z. u
Chapter 4 Manipulation of Potency
2 r) n# z; d. S. O Section A iPS Cells ) Y1 M0 Z7 H1 v$ b/ \+ R; ^) b# t
UNIT 4A.1 Human iPS Cell Derivation/Reprogramming ' ?5 z& S+ Y4 ^$ {6 c3 K. L! }
UNIT 4A.2 Generation and Characterization of Human Induced Pluripotent Stem Cells
5 V& B. X0 [/ U; }/ c$ P Section B Nuclear Transfer
, h) m' A- n0 _1 W8 VUNIT 4B.1 Heterokaryon‐Based Reprogramming for Pluripotency % D P+ z5 } V; U# w2 T
Chapter 5 Genetic Manipulation of Stem Cells 1 A' l# i) o9 d: [8 g
Section A Lineage Tracers in Stem Cells
- Y& H/ `; A5 u2 BUNIT 5A.1 Imaging Neural Stem Cell Fate in Mouse Model of Glioma ' w9 S+ p# o0 G/ w( }) {6 m
UNIT 5A.2 Functional Analysis of Adult Stem Cells Using Cre‐Mediated Lineage Tracing ) ^, U1 m3 @- h; r
Appendix 1 Useful Information . n1 {8 |% @0 J" ~2 o- K0 {
APPENDIX 1A Guidelines for the Conduct of Human Embryonic Stem Cell Research : ^/ D7 T0 v& p
APPENDIX 1B ISSCR Guidelines for the Clinical Translation of Stem Cells 8 y# q% d2 B% w% |3 S( y" N/ B( o
Appendix 2 Laboratory Equipment ( L1 ?1 B9 P5 ~" A- a$ ?3 v
APPENDIX 2 Standard Laboratory Equipment 2 \$ R# y! P& @
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