Hi all, need some help/ advises for a problem I encounter as mentioned in the title.
I am detecting iNOS (inducible nitric oxide synthase) protein from a murine cell line. After the western blot, I can see a clear and reproducible band. However, the band did not appeared at the expected location. the band is approximately 80kDA smaller that expected (expected molecular weight of the iNOS protein is 130kDA).
Does anyone know what may be the cause of the above phenomena? Is it possible that the protein ladder which I use is misleading?
Please kindly advise.
Thank you so much.
*attached are the picture of the blot membrane and the pre-stained protein ladder.
-lane 1 and 3: unstimulated cells sample
-lane 2 and 4: stimulated cells sample
-theoretically, stimulated cells will have higher iNOS expression than unstimulated one. the bands appreared showed the same trend. However, the bands appeared at different location than the expected location.