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Electrophoretic Mobility Shift Assay Forum EMSA Forum. Discuss the Electrophoretic Mobility Shift Assay protocols or technique and the assessment of DNA and RNA binding proteins here. Also post questions and troubleshooting about the shift assay.


Increased Gel Voltage to improve the Resolution?

Increased Gel Voltage to improve the Resolution? - Electrophoretic Mobility Shift Assay Forum

Increased Gel Voltage to improve the Resolution? - EMSA Forum. Discuss the Electrophoretic Mobility Shift Assay protocols or technique and the assessment of DNA and RNA binding proteins here. Also post questions and troubleshooting about the shift assay.


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Old 01-05-2010, 05:32 PM
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Default Increased Gel Voltage to improve the Resolution?



I want to solve two proteins that differ in 2 Kd (14 and 16kd respectively), but I cant get the resolution with my minigels (12%spacer+ 15%resolving): always appear one band instead of two separated bands. I have checked all the buffers and its OK (I think..) Now, re-reading the original paper (Schagger and von Jagow) says that the voltage AND the current must be fixed, for my gels the initial voltage must be 30V then 90V, AND the initial current may be as high as 80mA. I usually run at 30+90V and 400mA max current! So, I use Power (W) greater than the paper suggests and I still cant separate my proteins. But I dont understand, and correct me If I wrong: if you set the Voltage in the power supply, the Resistance offered by the buffer and gel will fix the current automatically, or at least that says the Ohm´s law if I remember well.. And a another question, can the product between the voltage and the current (Power) improve (or decrease) the resolution of the PAGE? If you know the right answer: thanks in advance
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Old 01-21-2010, 02:20 AM
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Default Re: Increased Gel Voltage to improve the Resolution?

Hi there,
usually the best way to get very sharp bands is constant current not voltage. Use stacking and resolving gels to optimize as well

let us know how you go
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Old 01-25-2010, 07:40 PM
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Default Re: Increased Gel Voltage to improve the Resolution?

Try 18% gel. Run at 100V flat.
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