I am doing western blot with brain protein samples. I loaded replicates of 5 samples in 4-12% Bis Tris gel and cut it in half.
I found that sample 2 and 3 are streaky at replicate #1, but sample 2 was still streaky but sample 3 was not streaky anymore in replicate #2.
I loaded them all at the same time with the same pipette. While gel was running, all the other samples were migrating with flat line shape but the streaky samples were migrating with thin triangle shape, so that we already knew those samples will be streaky like something is pulling them down the gel.
Does anyone know what could cause streaky bands on SDS-PAGE?
Hello Ky Lee,
several factors can contribute to streaking in SDS polyacrylamide gels:
- protein overloading (I would not load more than 30 microgram per lane)
- voltage/current of the electrophoresis is set too high
- inhomogenities of the electric field due to a faulty electrophoresis apparatus
- impurities in the sample (lipids, insoluble matter, etc.)
- not properly rinsing to gel pockets prior to sample loading (this removes unpolymerized acrylamide from the pocket)
Best regards,
Christian
Hi Ky Lee,
I would re-spin samples. Most probably there are some impurities in the sample as suggested by Christian.
Best,
Lena
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