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( G. X, ^. T4 O4 |Current Protocols in Stem Cell Biology6 E/ M, M& Y; Q; U d c2 `
Last Update: July 07, 2009
% Y4 T' s3 x. V5 W S# F5 q+ VPrint ISSN: 1941-7322; z1 d0 @2 p1 c
Online ISSN: 1938-8969 _5 O3 l7 B( o1 Y, }9 Y$ W) B0 {
Table of Contents+ m4 w/ R" _: R) R4 H q
Foreword / f# t/ v- @2 g% @9 ]) d
Preface ' B5 ~( x3 F" A) f1 A6 x2 [
Chapter 1 Embryonic and Extraembryonic Stem Cells
; M- y" p8 v* ^" K b- J Section A Isolation of Embryonic Stem Cells
7 V1 |, [7 G: v. GIsolation of Embryonic Stem Cells m$ A1 V2 }" t, N
UNIT 1A.1 Derivation and Characterization of Nonhuman Primate Embryonic Stem Cells % O; C; f* \9 V+ r! p
UNIT 1A.2 Derivation of hESC from Intact Blastocysts
! u1 ]' c! f ?' tUNIT 1A.3 Reprogramming Primordial Germ Cells (PGC) to Embryonic Germ (EG) Cells " o* J; d9 e+ d7 f; p, e
UNIT 1A.4 Derivation and Propagation of hESC Under a Therapeutic Environment
: c4 b- X+ P4 d( M& D! ^ Section B Characterization of Embryonic Stem Cells 4 t& p% z$ i/ |: |% L, b! _% L
UNIT 1B.1 Proteomic Analysis of Pluripotent Stem Cells 2 t1 t t ?- D- c
UNIT 1B.2 Gene Expression Analysis of RNA Purified from Embryonic Stem Cells and Embryoid Body–Derived Cells Using a High‐Throughput Microarray Platform
% Z# d! k# G; dUNIT 1B.3 Phenotypic Analysis of Human Embryonic Stem Cells
% _$ q# q" t# M8 J3 }) T( g+ I& BUNIT 1B.4 Isolation of Human Embryonic Stem Cell–Derived Teratomas for the Assessment of Pluripotency
1 _; X" c9 Z: c7 v" j# c, RUNIT 1B.5 Tandem Affinity Purification of Protein Complexes in Mouse Embryonic Stem Cells Using In Vivo Biotinylation
- s$ p- k5 Z. J' k R6 ~3 O: C Section C Culture and Maintenance of Undifferentiated Embryonic Stem Cells
0 U8 O/ S# |5 Z8 FCulture and Maintenance of Undifferentiated Embryonic Stem Cells
2 {* p! K' l3 A( ^UNIT 1C.1 Expansion of Human Embryonic Stem Cells In Vitro ) f" q% f8 e& k- @
UNIT 1C.2 Defined, Feeder‐Independent Medium for Human Embryonic Stem Cell Culture
. O- _3 {% W1 c- A& v5 f; PUNIT 1C.3 Isolation and Propagation of Mouse Embryonic Fibroblasts and Preparation of Mouse Embryonic Feeder Layer Cells ; Q: h, ?: N j9 q6 @
UNIT 1C.4 Culture of Mouse Embryonic Stem Cells 0 t; d+ q1 G6 M+ D; F6 P
UNIT 1C.5 Preparation of Autogenic Human Feeder Cells for Growth of Human Embryonic Stem Cells 0 [+ x" w+ k# `# b0 f: @
UNIT 1C.6 Isolation of Human Placental Fibroblasts
7 g; ^ b& V9 l% UUNIT 1C.7 Derivation of Human Skin Fibroblast Lines for Feeder Cells of Human Embryonic Stem Cells - Z2 j" l) @% I; m1 g
UNIT 1C.8 Cryopreservation of Dissociated Human Embryonic Stem Cells in the Presence of ROCK Inhibitor
" I. K2 l% H- A; X9 M Section D Germ Layer Induction/Differentiation of Embryonic Stem Cells " P! C r3 _; C) Z1 E% D+ n x$ D
UNIT 1D.1 Germ Layer Induction in ESC—Following the Vertebrate Roadmap
' x5 _( U2 Y# UUNIT 1D.2 Formation and Hematopoietic Differentiation of Human Embryoid Bodies by Suspension and Hanging Drop Cultures ) ]" K0 a1 [, A/ m) f% u
UNIT 1D.3 Directed Differentiation of Human Embryonic Stem Cells as Spin Embryoid Bodies and a Description of the Hematopoietic Blast Colony Forming Assay / e) f3 A# u! w( a+ K5 k8 o+ Q$ k; e
UNIT 1D.4 Differentiation of Human Embryonic Stem Cells in Adherent and in Chemically Defined Culture Conditions [; E' I( |. J: {5 k3 C1 {
UNIT 1D.5 Isolation and Differentiation of Xenopus Animal Cap Cells
* ]' k/ m$ w, N+ K# H Section E Extraembryonic Lineages
# a' }0 s0 T* ~6 P9 @Isolation of Stem Cells from Extraembryonic Tissues # L6 L/ V$ O1 I6 \& h7 l7 v: d
UNIT 1E.1 Isolation of Human Placenta‐Derived Multipotent Cells and In Vitro Differentiation into Hepatocyte‐Like Cells ! @6 T; O1 ?& {. y- z: y9 [
UNIT 1E.2 Isolation of Mesenchymal Stem Cells from Amniotic Fluid and Placenta
/ m; t# f2 s( ?! }8 J$ L4 HUNIT 1E.3 Isolation of Amniotic Epithelial Stem Cells
, P U9 f% W! f7 GUNIT 1E.4 Isolation and Manipulation of Mouse Trophoblast Stem Cells
8 u5 G/ \* b* e$ S2 l6 |# [ Section F Mesodermal Lineages + x+ }' K* j: P2 G
UNIT 1F.1 Differentiation of Embryonic Stem Cells into Cartilage Cells
& W. H0 }0 n1 EUNIT 1F.2 Differentiation of Human Embryonic Stem Cells to Cardiomyocytes by Coculture with Endoderm in Serum‐Free Medium
( M# C* |! m: oUNIT 1F.3 Isolation of Hematopoietic Stem Cells from Mouse Embryonic Stem Cells
; ]( F* E" y5 U" p1 [7 d5 bUNIT 1F.4 Differentiation of Mouse Embryonic Stem Cells into Blood
1 p0 u8 h3 f& |: ~% x" d6 C, |0 O9 nUNIT 1F.5 Endothelial Differentiation of Embryonic Stem Cells
( _7 }1 c P( L. cUNIT 1F.6 Hematopoietic Differentiation of Human Embryonic Stem Cells by Cocultivation with Stromal Layers
* |* n c% f# T2 ZUNIT 1F.7 TLX1 (HOX11) Immortalization of Embryonic Stem Cell–Derived and Primary Murine Hematopoietic Progenitors 3 o6 t1 a9 |( x" ?% c N/ D# Q; w
UNIT 1F.8 Differentiation of Multipotent Mesenchymal Precursors and Skeletal Myoblasts from Human Embryonic Stem Cells w. h2 c6 s( X, X+ e
Section G Endodermal Lineages
( J, M8 }3 k$ w+ I. uUNIT 1G.1 The Differentiation of Distal Lung Epithelium from Embryonic Stem Cells / y" z" \% C. L2 I
UNIT 1G.2 Pancreas Differentiation of Mouse ES Cells , n3 i- r1 v+ Z8 |" j& L# R6 A7 ?- K
UNIT 1G.3 Differentiation of Embryonic Stem Cells into Anterior Definitive Endoderm 2 L1 c: U3 ^! ]- g: w& K' @
Section H Ectodermal Lineages * c" _6 z: X0 t8 Z l Z- O
UNIT 1H.1 Differentiation of Mouse Embryonic Stem Cells to Spinal Motor Neurons
! |* U. d7 D) I; o2 U3 w Chapter 2 Somatic Stem Cells ! Q8 {' w& D( J0 H4 x/ |$ g9 T( u
Section A Hematopoietic Stem Cells % A q0 S/ o# W$ U6 j7 r& Y% G
Hematopoietic Stem Cells 9 {% c# R; [+ Q
UNIT 2A.1 Isolation of Mononuclear Cells from Human Cord Blood by Ficoll‐Paque Density Gradient 8 F9 q T- g- `
UNIT 2A.2 Isolation of Hematopoietic Stem Cells from Human Cord Blood % S( \# J% R0 V X% `9 O
UNIT 2A.3 Isolation of Mesenchymal Stem Cells from Human Cord Blood 8 K+ {& v$ B2 k' E" x& ]: A x' r
UNIT 2A.4 Isolation and Assessment of Long‐Term Reconstituting Hematopoietic Stem Cells from Adult Mouse Bone Marrow . W: S" b* S" V* c5 {
UNIT 2A.5 Analysis of the Hematopoietic Stem Cell Niche
1 y2 o! ~. D+ o5 ]: f( {( NUNIT 2A.6 Analysis and Manipulation of Hematopoietic Progenitor and Stem Cells from Murine Embryonic Tissues
; F+ a& F1 I, J. g9 d! f' HUNIT 2A.7 High Level In Vitro Expansion of Murine Hematopoietic Stem Cells
0 P" J k5 h4 ?# W3 Y' k; n- k# HUNIT 2A.8 Isolation and Visualization of Mouse Placental Hematopoietic Stem Cells ; G' C- }. \8 s
Section B Non‐Hematopoietic Bone Marrow‐Derived Stem Cells " H; q; h2 E7 l9 h( O
UNIT 2B.1 Isolation and Characterization of Mesoangioblasts from Mouse, Dog, and Human Tissues 8 K9 J$ O, s3 e! q- R
UNIT 2B.2 Purification and Culture of Human Blood Vessel–Associated Progenitor Cells ( @5 G" t V, |- d
UNIT 2B.3 Isolation, Culture, and Differentiation Potential of Mouse Marrow Stromal Cells 5 F& K) j' Q4 |! p
Section C Cardiovascular Stem Cells
% f2 a% f( Q1 Y9 ?3 QUNIT 2C.1 Isolation and Characterization of Endothelial Progenitor Cells from Human Blood
# j+ w0 V0 j3 Q; u6 HUNIT 2C.2 Derivation of Epicardium‐Derived Progenitor Cells (EPDCs) from Adult Epicardium - ^. b% x& x' ]6 N& p; X
Section D Neural Stem Cells ' E* Z c' Z) Q$ ?8 V! `
UNIT 2D.1 Generation of Cultured Oligodendrocyte Progenitor Cells from Rat Neonatal Brains ! ^ Y/ V* f4 y; M4 ^" B7 S
UNIT 2D.2 Isolating, Expanding, and Infecting Human and Rodent Fetal Neural Progenitor Cells ( V5 x0 O: l$ D* E8 A' N. ^. r$ x
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 * t' X; I" N% {, }; J3 r
Chapter 3 Cancer Stem Cells
+ n3 ^: c6 J0 H# A2 hUNIT 3.1 Colon Cancer Stem Cells " L0 p7 G) w6 \2 Z1 W! _
UNIT 3.2 In Vivo Evaluation of Leukemic Stem Cells through the Xenotransplantation Model
4 [# I4 x. c, L. C0 Y( S Chapter 4 Manipulation of Potency
* c2 c$ f% L+ w# l# l, h# U Section A iPS Cells
. D& ~; o, |1 e( BUNIT 4A.1 Human iPS Cell Derivation/Reprogramming 3 R9 k; Y. `: R7 y( R5 S3 U
UNIT 4A.2 Generation and Characterization of Human Induced Pluripotent Stem Cells
4 t @7 e9 _9 w3 v* C9 i Section B Nuclear Transfer
, U Q2 [, t! N: k2 F' O4 ^$ ^4 ^UNIT 4B.1 Heterokaryon‐Based Reprogramming for Pluripotency
/ h0 R9 x9 A$ f& `& F% V3 l+ [6 W Chapter 5 Genetic Manipulation of Stem Cells
! W5 g g' }, I9 V- l Section A Lineage Tracers in Stem Cells 2 L3 h$ _- N0 P
UNIT 5A.1 Imaging Neural Stem Cell Fate in Mouse Model of Glioma
: t1 h7 f9 y) w2 rUNIT 5A.2 Functional Analysis of Adult Stem Cells Using Cre‐Mediated Lineage Tracing
, {3 v1 M6 t/ n- ]1 V ?1 n2 l8 N Appendix 1 Useful Information 9 s D: ]+ V" [* b8 g0 G% z
APPENDIX 1A Guidelines for the Conduct of Human Embryonic Stem Cell Research
2 U4 {- S- O$ X0 X! CAPPENDIX 1B ISSCR Guidelines for the Clinical Translation of Stem Cells 6 _0 d2 S; ~1 I2 B
Appendix 2 Laboratory Equipment
0 P1 \" c( Y; Q4 E3 u: n D5 uAPPENDIX 2 Standard Laboratory Equipment
4 Q4 N5 L2 M1 e/ U( Z7 x: @' l先谢谢啦! |
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