I am seeing a lot of background staining on pIMAGO kit for phosphorylated proteins on membrane. I am using PVDF membrane . All reagents were freshly prepared with milliQ water etc. Does anybody know which membrane gives the lowest autofluorescence ?
hi
1- It seems natural. The kit generally recognized as phosphate groups( the cell lysate full of phosphate groups) .
2- you should test 1:5000 dilution from pIMAGO reagent.
3- decrease loading protein lysate amount.
4- one sample lysate used as negative control ( lysate with out phosphatase inhibitor )
good luck
thanks Reza
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