"Hello! My friend. Stay awhile and listen!"

You may able to save soul of a poor graduate student.

I have trouble native-PAGE.

Here's my test condition.

Rice soluble protein, extracted by phosphate buffer, with liquid nitrogen, mortal and pestle, 12,000 g 10 min centrifuged.

I used 7% and 10% for seperation, 4% for stacking(10 ~13 mm) acrylamid gel with Tris buffer.

Loading buffer was tris-glycine buffer.

40 ug/ 16ul protein was loaded in the well.

Four gels(2 of 0.75mm, 2 of 1.0 mm)with 2 tanks(bio-rad) was run 30 mA, 20 min for stacking, 60 mA, 60 min for seperating.

And did some antioxidant color stainning for CAT, POD, SOD

One day I could get a perfect results, but usually got a strange bands.

I was NEVER seen these shape of bands in the SDS-PAGE, so I don't even think about what's wrong in my native PAGE.

Do I need to filter with miracloth?

Would you help me why my bands were going to be shrunken and dragged?

Thanks

고맙습니다.

P.S

Thanks for all of advices. By the way in my case the problem was solved partially to centrifuge samples 3 times. Also the casting of stacking gel could be problem.