|
Gel shift assays, or electrophoretic mobility shift assays (EMSA),
provide a simple method to study DNA-protein interactions. Existing
mobility shift assay protocols can be easily converted to infrared
fluorescent assays by replacing the existing DNA oligonucleotides
with IRDye® infrared dye end-labeled oligonucleotides. Binding
and electrophoresis conditions are the same as any other EMSA
detection method.
A DNA oligonucleotide end-labeled with IRDye 700 infrared dye
is a good substrate for protein binding. Using the Odyssey Infrared
Imaging System, IRDye infrared dye labeled DNA detection is linear
within a 50-fold dilution range from 9.1 fmol to 0.18 fmol. Infrared
assays can be completed in less than two hours with no gel transfer
or film exposure. The gel doesn’t even have to be removed
from the glass plates for imaging. Plus, the gel can be placed
back into the electrophoresis unit and run longer, if necessary.
| Advantages of Infrared |
| Infrared |
Radioisotope |
| Easy access and disposal |
Regulatory procedures
Disposal limitations |
Dye is stable for a long time
|
Short half-life of the label |
| Non-hazardous |
Hazardous |
| Gel (glass plates) can be easily
imaged on the Odyssey |
Lengthy incubations with autoradiographic
film |
| Detection of the probe is rapid |
Time-consuming and inconvenient |
| Gel can be replaced back and run
longer |
Not possible |
IRDye 700 infrared dye-labeled EMSA oligonucleotides are supplied
as 25 μl of 50 nM double-stranded DNA
Please note: Oligos with an * require up to two weeks for processing and delivery.
| Part # | Description | Downloads | Order Online |
| 829-07910 | EMSA Buffer Kit for the Odyssey
| description
msds
protocol
| $ 195.00
 |
| 829-07925 | EMSA Oligo, AP-1 (IRDye 700)
| msds
protocol
| $ 50.00
|
| 829-07933 | EMSA Oligo, ARE (Androgen Receptor) (IRDye 700)*
| description
msds
protocol
| $ 50.00
|
| 829-07923 | EMSA Oligo, CREB (IRDye 700)*
| msds
protocol
| $ 50.00
|
| 829-07927 | EMSA Oligo, E2F (IRDye 700)*
| msds
protocol
| $ 50.00
|
| 829-07929 | EMSA Oligo, HIF-1 (IRDye 700)
| msds
protocol
| $ 50.00
|
| 829-07924 | EMSA Oligo, NFkB (IRDye 700)
| msds
protocol
| $ 50.00
|
| 829-07930 | EMSA Oligo, OCT-1 (IRDye 700)*
| msds
protocol
| $ 50.00
|
| 829-07931 | EMSA Oligo, PAX-5 (IRDye 700)*
| msds
protocol
| $ 50.00
|
| 829-07926 | EMSA Oligo, Sp-1 (IRDye 700)
| msds
protocol
| $ 50.00
|
| 829-07922 | EMSA Oligo, STAT3 (IRDye 700)*
| msds
protocol
| $ 50.00
|
| Part # | Description | Downloads | Order Online |
| 829-07910 | EMSA Buffer Kit for the Odyssey
| description
msds
protocol
| $ 195.00
|
|
Tube |
Reagent (Reagents for 100 EMSA Reactions) |
Volume |
1 |
10 X Binding Buffer (100mM Tris, 500mM KCl, 10mM DTT; pH
7.5) |
500ul |
2 |
25mM DTT/ 2.5% Tween-20 |
500ul |
3 |
Poly (dI-dC) (1ug/ul) in 10mM Tris, 1 mM EDTA, pH 7.5 |
125ul |
4 |
Sheared Salmon Sperm DNA (0.5 ug/ul) in 10mM Tris, 1 mM
EDTA, pH 7.5 |
125ul |
5 |
50% Glycerol |
500ul |
6 |
1% NP-40 |
500ul |
7 |
1M KCl |
500ul |
8 |
100mM MgCl2 |
500ul |
9 |
200mM EDTA, pH 8 |
500ul |
10 |
10X Orange Loading Dye |
500ul |
| Storage: -20°C |
Shipping information: U.S.A.: Same-day shipping (Federal Express overnight) available on orders received before 12:00 noon CST. Canada: Allow 3-5 days for shipments. Germany & UK: Contact LI-COR GmbH. International: Contact a LI-COR distributor in your area. All shipments subject to availability. |
