I have run an SDS PAGE and after Silver-staining, I am getting extra thin lanes (Vertical) bifurcated from main lane. What could be the possible expalnations for these thin lanes??
Hello, the most possible cause of bifurcation of the lane can be the overloading. Moreover. overflowing of the samples cause the samples to be stuck over the outer well walls and thus, forms another lane. Silver staining being very sensitive leads to its easy visualization as another lane or bifurcation.
Hope this helps!
Hello Sandeep,
Trying to judge without knowledge of the kind of protein sample you are running, I think that splitting of lanes can occur if the quantity of protein loaded per well is too much. Also, using high run voltage in addition to large sample volumes/protein load may cause deformations in lanes. Another possibility could be how the gel was prepared especially how the wells were......sometimes it is not clearly visible but an intermediate ridge may make one well...two and a large sample just spills into the other sub-well.
Try to remake the gels and check the wells. If you use commercial gels, try to load a smaller sample both in terms of volume and if possible protein concentration.
Good Luck
Hello, the most possible cause of bifurcation of the lane can be the overloading. Moreover. overflowing of the samples cause the samples to be stuck over the outer well walls and thus, forms another lane. Silver staining being very sensitive leads to its easy visualization as another lane or bifurcation.
Hope this helps!
Try loading lesser amounts of sample in each well in your next run. Also, do not leave any gap wells in between as these can cause sample diffusion and lane widening. If nothing else, load the gap wells with your loading dye. Also, do not go for too high a run voltage.
Thank you everyone for valuable suggestions...
Sample type: Serum
Protein conc.: 6 ug
Sample volume: ~16-18 ul
Inhomogeneities in your mixing of the gel might also do this, if you mix your own. You can also buy precast gels and then blame it on the manufacturer if things go wrong. If you do use precast gels, then sample overload is more likely.
Hi Sandeep.
I would suggest you to load ten microlitre of sample in each well and see if there is any improvement. Also as I mentioned earlier, do not leave any empty wells. Fill them up with loading dye to prevent lane widening and diffusion. Lets hope your gel is fine.
Best
Aditya
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