PDF电子书：Western Blotting Handbook and Troubleshooting Guide(version 2)

dahui

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Introduction  1# Y2 x, u7 L/ K: H6 h1 n' T
Western Blotting Overview  2-3& |' ~9 c* T$ g' `5 n/ ~
Step 1 – SDS-PAGE . U2 L  ?( f) j7 ]6 l# J
  Thermo Scientifc Precise Protein Gels  4
5 \  H* q1 b6 t' {' g, k5 Q) p  Molecular Weight Markers  5-6
; P9 o; c4 t  q4 S/ C: @Step 2 – Electro-Transfer * U% }) a0 U) G% \9 V: y
  Thermo Scientifc Pierce Fast Transfer System  7 ; q5 s6 T, o7 v' |$ Y9 J
  Transfer Buffers  8 6 ]! D- y1 ~+ q+ b5 ~3 u2 ]. Z
  Filter Paper for Blotting  8 . C. B) Z$ G" U' `9 H+ M$ \" M1 I
  PVDF and Nitrocellulose Membranes  9
9 \3 d7 t# S  X: e/ C1 o) `. p, [  Thermo Scientifc Pierce Protein Stains   9  & o: A5 |- @% I, u, o7 p# ]* N+ G0 H; F
   for Membranes     y$ l/ W( ]- T8 Z8 _' M8 s
  Antibody Extender Solution NC  10
& m* ~- Y* e* Q: ~' @/ q; Z  Western Blot Signal Enhancer  11" J$ a1 z. f& j6 i3 A) s7 ]1 s2 U' N
Step 3 – Blocking
+ K* z9 W, U* `& a; |  Introduction  12
( W/ O$ i2 E9 c+ H, E  w" s; h  Blocking of Nonspecifc Binding Sites on  12
& i7 f6 T: S% P, y# L# E   Transfer Membranes   , B! i+ S% N' l) I0 _
  Blocking Buffer Optimization  12-13
: B8 Z* ]; A* b( a" @- p4 l1 A4 l  Blocking Buffers  13-163 H. `, o* t/ b0 I
Step 4 – Formulate Wash Buffers 6 J- h. d0 c* |( I
  Washing the Membrane  17
( }: F( ^/ J5 G  Wash Buffers  17; W- c3 |4 R& d9 W4 z% o! V+ C
Step 5 – Detection Reagents
+ J) K7 U0 `+ s0 ^/ S) g- @4 m  Validated Primary Antibodies  18-19 9 L: s- J% i4 Z/ |1 [* U$ P
  Affnity-purifed Antibodies  20 ; m+ b7 m! u6 ?- {1 g) J9 i2 D
  Stabilized HRP Conjugates  21
/ m: w$ L/ s; M# D( M  i4 r: _! r  Thermo Scientifc DyLight   22
6 B" x8 c, J# k7 j   Fluor Conjugates   
/ A% d0 z# k( ^" i0 S+ p  Conjugate Stabilizer Solutions  22
6 _5 W* j! u0 W: E2 d8 c$ ?  Thermo Scientifc DyLight-labeled Highly  23
" W: Q& j. j) g* c   Cross-Adsorbed Secondary Antibodies 7 J% T9 U" j0 w) |
  Antibody Storage and Stabilizer Solutions  24 % [, F" t, H5 I$ S
  Secondary Antibody Ordering Table  25-26  8 _4 ]4 z1 x5 q6 q
  Thermo Scientifc Clean-Blot IP  28-29
# t! r0 J" r; f' S5 @   Detection Reagents   % u0 L9 l+ q: U' |# e7 K3 |$ p! U
  Protein A, G, A/G and L Conjugates  30 ! g$ J/ b( }1 P' |) S3 P9 g
  Thermo Scientifc NeutrAvidin, Streptavidin   31-33 & N. U$ A7 k5 i  [9 l1 e1 W+ i' g
   and Avidin Conjugates
# }5 k- i' g" T& W: `: R7 q' ?6 uStep 6 – Enzyme Substrates " N8 ]1 a! C2 u6 s3 j
Chromogenic Substrates  34-35 ! u* [( L6 r- s- q. p1 z- B( y
Thermo Scientifc Chemiluminescent Substrates  36-43 7 ~( i' M' X- R) v) ^9 P6 |
  Pierce ECL Substrate  37
4 _" ]3 v6 m1 T! @; e2 e" x8 m  SuperSignal Chemiluminescent   38-42
5 m' L" ~1 g. J! }   Substrates and Kits   
3 E- c+ I8 w# D/ y0 ?  Lumi-Phos™
) O5 r4 i" G/ F6 h Chemiluminescent Substrate  43
. ]$ {, S2 N$ X1 L! x& H+ w+ _  Quick Reference Substrate Guide  43 ! F, G4 g0 t5 N1 b6 P  _) a4 S
Fast Western Blotting Kit  44
- D  F0 n1 n+ j0 a5 d9 aSpecialized Western Blotting Kits  45-48
/ s* B2 F: X) Y0 E) j" x( ~6 z  Thermo SuperSignal West Pico HisProbe Kit  45 0 ?$ N' ~) k% A: U$ o: ~! f4 F: ?
  Pierce O-GLcNAc Western Blotting Detection Kit  46
0 M. V3 q. y5 j( f  Thermo Scientifc DyLight 549/649  46-47  % C7 d& Z! c: \! c6 F. S
   Western Blotting Kit   5 F" [( j8 }. I/ c+ v$ o( f# \
  Thermo Scientifc DyLight 680/800 Near Infrared  47 " A; |! h7 W, [" v
   Western Blotting Kit   4 ~; d  h6 V) Y5 q) R! r
  Thermo Scientifc Active GTPase Pull-Down  48
9 l' N3 \7 T3 N% K4 I9 ^9 k   and Detection Kits   
; _: m, k0 V) W" t3 FFar-Western Blotting  49
& V; ^) |3 J# l3 M4 VIn-Gel Western Detection  50-51 $ f. c* R# g: {2 o  z. p6 w
  Thermo Scientifc Pierce In-Gel   51
% u" K' T4 S! @6 L1 n2 W$ O   Chemiluminescent Detection Kits3 p# W: r$ Z0 ?
Step 7 – Film
  P$ ^/ I& W6 l$ b6 T8 J  Thermo Scientifc CL-XPosure Film  52% T( O$ e$ O2 @7 s4 x5 J
Step 8 – Stripping Buffer
( e7 g1 |& C/ e/ k4 G- COptimizing the Signal-to-Noise Ratio  53-58 ) W  Z" m4 A6 v6 V4 J% n! e* Y4 C7 I
  Protocol for Stripping an Immunoblot  54
8 C  A0 @6 d) n0 \5 X  Thermo Scientifc Restore  55
, N7 ?9 k6 J% n8 K* Q   Western Blot Stripping Buffers
5 L+ P+ y) ]9 R! e% N  Thermo Scientifc Restore PLUS  56
( u0 T6 j( a" N   Western Blot Stripping Buffers
- |# [+ j- K/ T) V. k# e  Thermo Scientifc Pierce   57-58 $ k6 e! G5 S& Z4 }( [
   Background Eliminator
; i/ D0 S/ {6 _' G" |Troubleshooting Guide  59-69
" _" c' j6 R# p% t3 D$ Q& |  Blotting with Chemiluminescence  59
+ [. I3 i! H/ h  Optimizing Antibody Concentration  60-63 * M$ s4 z% B  H8 S) \
  Problem Guide  64-67
& _1 P% i$ O% Q4 t  Full-Length Western Blotting Protocol Using   68-69 6 o" ~- N& Y$ W6 `/ M  j
   Chemiluminescent Substrates
' A- ?0 a/ q5 nRecommended Reading  70
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一枝独秀

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jiandong2001

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军医

谢谢分享！

深海寂寞鱼

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白衣小彩

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sunyucheng

看看

daviddow

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mominglily

谢谢

qlwj

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