When I was doing a SDS-PAGE upto 25kD the bands of the protein ladder got resolved properly , below the 25kD the 20, 15 and 10kD bands did not resolved and the tracking dye of the samples was moving along with the 25kD band of the ladder. What are the possible reasons for this ?
Hi
What percentage of the gel are you running? If you running a low percentage gel then the resulting large pore size can result into what you are observing. Are you using BIORAD's precision plus ladder?
Vikrant Mehta I am using 7.5% resolving gel . Earlier also, I had used this percentage of gel but it never happened like this . Yes, I am using BIORAD's precision plus ladder. Can you tell me if the samples above 25kD has been resolved properly or not ?
Hi Deeptimayee,
For protein sizes you are using, acrylamide percentages 14-16% could be adequate to a better definition.
In a 7.5% gel, i would not expect a band lower than 20 kD to show up in my resolving gel. I have run 8% gel and believe me I was able to see the ladder up to 37kD only. (Please see attached images as well). It's rather strange to me how would you get to see bands lower than 25kD as you have mentioned.
Do you have any representative images that could give clear idea? Please go through and check the recipe again that you used to prepare the gel in both the cases.
Deeptimayee Guru I appreciate that. Please let me know if you were able to solve this or any new observations in this regard.
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