Verticals streaks in kinase assay gels - (Dec/21/2005 )

Hi everyboby,

I am kind of desperate. I did 5 experiments (phosphorylation of myelin basic protein by a protein immunoprecipitated from a cell lysate) and each time got the same pattern: vertical streaks, of random size and distance from each other (not same distance as the comb lanes). But the stained molecular weight marker look ok! Any ideas? I use a Mini Protean 3 form Bio-rad and run a 20% gel.

Thank you!

did you load too much of proteins?
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Thank you for this suggestion

I used myelin basic protein at a concentration of 0,5mg/ml, so I should have 40ug per lane. The problem is that I did this experiment before with this same concentration and it never was a problem. It can't be a mistake in the dilution of MBP because I tried 2 different bottles.

Is it possible to IP too much proteins? Do those immunoprecipited proteins end up on the gel too? If so I should see a difference between the negative control (IP with normal mouse IgG) and the lanes using anti-HA antibody but I don't.

Thanks again