Sample Application Isoelectric Focusing
We got the following question:
I reversed the position of the gel … In this experiment everything went right. The bands were extremely clear and the bands from the marker were straight … I would like to ask if the result obtained is the effect of protein migration from higher to lower ampholyte concentrations.
SERVA TechServ replied:
In IEF, it doesn’t matter where you apply your sample. The protein always runs to the isoelectric point. The main factor is the mobility and solubility of the protein. These are influenced by many factors, e.g. Buffer, pH value, hydration shell and much more. Perhaps your protein migrates better because the pH value / the ampholytes are more suitable at the new application point and the protein therefore migrates better into the gel and has better mobility …