ECL plex Product Specialist 1/
Western Blot 2/
Western Blotting- PI 3/
Western Blotting 1. 2 1 2. 3. 4. Y 3 4 5. 4/
1. SDS- (SDS-PAGE) SDS ( β- ) --SDS 5/
SDS-PAGE - - - (Tris-HCL ph 6.8) glycine/chloride (Tris -HCL ph8.8) Tris-glycine ph 8.3 + + + 6/
2. : ( - ) 7/
+ - l 8/
3. CyDye Y Y 1 Y Y Y Y 2 Y Y Y Y 9/
HPR AP 10 /
Chemiluminescence Fluorescence Chemifluorescence Radioactivity ( 35 S or 125 I) 11 /
4. 12 /
ECL ECL Plus ECL Advance ECL Plex 13 /
ECL HRP Peracid HRP Luminol + Proprietary Enhancer Hyperfilm ECL / CCD 14 /
ECL order information RPN 2125 ECL and ECL Plus blocking reagent for 2000cm 2 15 /
ECL vs. ECL 125 I DAB NBT/BCIP a b c d IgG Hybond TM ECL, : a), ECL, Hyperfilm TM ECL 15 b) 125I-anti-mouse (0.6μCi/ml), 18 hr c), DAB 20 min d), NBT/BCIP 20 min 19 /
ECL Plus -HRP ECL Plus ( 503nm) /CCD ( 430nm) 20 /
ECL plus order information RPN 2124 ECL Plus Western blotting Reagent Pack anti-mouse IgG horseradish peroxidase conjugate, 100 µl; anti-rabbit IgG horseradish peroxidase conjugate, 100 µl; ECL membrane blocking agent, 5 g. Sufficient for at least 10 mini-blots. RPN 2125 ECL and ECL Plus blocking reagent for 2000cm2 21 /
ECL Advance ECL Advance -HRP ECL Advance HRP Hyperfilm ECL / CCD 22 /
ECL Advance order information RPN 2135 ECL Advance Western Blotting Detection kit for 1000cm 2 23 /
Sensitivity Comparisons Using Optimised Antibody Concentrations: Nitrocellulose Membrane: Hybond ECL ECL Advance ECL Plus Pierce West Femto Primary 1:50,000 Secondary 1:200,000 Primary 1:5,000 Secondary 1:10,000 Primary 1:5,000 Secondary 1:100,000 PVDF Membrane: Hybond-P ECL Advance ECL Plus Pierce West Femto Primary 1:50,000 Secondary 1:200,000 Primary 1:5,000 Secondary 1:10,000 Primary 1:5,000 Secondary 1:100,000 24 /
ECL Western 25 /
ECL Plex Cy5 Cy5 Cy3 Cy3 CyDye = 1 = 2 26 /
Cy5 Cy3 27 /
ECL Plex with Cy3 and Cy5 : nm= Typhoon 9410 (nm) (nm). Cy3 550 570 532 580 BP 30 Cy5 649 670 633 670 BP 30 28 /
ECL Plex: Typhoon CyDye 29 /
ECL Plex : ( ) 30 /
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ECL ECL Plex, ECL Advance, ECL Plus, ECL ECL Plex 1:750 1:2500 Hybond-ECL (R 2 ) 0.998 (pg) 3.6 1.2 ECL Advance 1:100000 1:400000 Hybond-ECL 0.978 2.4 0.6 ECL Plus 1:10000 1:100000 Hybond-ECL 0.997 2.4 4.9 ECL 1:1500 1:10000 Hybond-ECL 0.961 1.5 9.8 A model system was used to compare the four systems to each other with minimal variability ECL Plus Fluorescent 1:10000 1:100000 Hybond-ECL 0.994 2.1 39 33 /
ECL HRP ECL Advance ECL Plus ECL ECL Plex (pg ) * ~1 pg ~5 pg ~10 pg ~1 pg Up to 1/100,000 Up to 1/500,000 Up to 1/20,000 Up to 1/5,000 1/100-1/5000 Up to 1/200,000 Up to 1/15,000 1/1250-1/4000 4-6 hrs 24-48 hrs 1-2 hrs Up to 3 months Yes Yes Yes (No) Hybond P or Hybond ECL Hybond P for ChemiLum Hybond ECL for ChemiFluor Hybond ECL Hybond ECL or LFP membrane Hyperfilm ECL / CCD Fluorescent Scanner / Hyperfilm ECL / CCD Hyperfilm ECL / CCD Fluorescent Scanner, &. *in model system 34 /
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1. 2.Hybond 3.
Tank Transfer TE22, TE62 Semi-Wet Transfer minive blot module Semi-Dry Transfer (ECL Semi-Dry Blotters, TE70/77 PWR) 38 /
Tank Transfer TE22 TE62 39 /
Semi-Wet Transfer Mini-VE Blot Module 40 /
ECL Semi-Dry Blotters TE70/77 TE70/77 PWR TE 70: 14 x 16 cm TE 77: 21 x 26 cm 41 /
Features and Benefits 42 /
ECL Multiprobe and ECL Multiprobe XL Decaprobe Incubation Manifold OLD 43 /
Hybond ECL 100% ECL 44 /
Hybond-P PVDF-polyvinylidene difluoride ECL Plus (lumi.) ECL Advance 45 /
Hybond-LFP Low Fluorescent PVDF-polyvinylidene difluoride ECL Plex PVDF 46 / MetOH Water Transfer buffer
Molecular Weight Markers Rainbow Markers ECL DualVue RPN810 M r (Da) 250 000 160 000 105 000 75 000 50 000 35 000 30 000 25 000 15 000 10 000 separation on a 12% SDSpolyacrylamide gel RPN756: RPN755: RPN800: MW Range High Low Full Range Size Range (kda) 14.3-220 2.3-46 10.0 250 47 /
ECL Plex On Typhoon (12% gel) Cy5 Cy3 250 k 160 k 105 k 75 k 50 k 35 k 30 k 25 k 15 k 48 /
Questions ECL ECL Plus ECL Advance ECL Plex ECL plex 3 49 /
800-810-9118 www.gelifesciences.com http://www.ebiotrade.com/ge/a KTAclub7/index.html 50 /