Which of the following is not associated with isoelectric focusing?

• A. Continuous pH gradient
• B. Migration of proteins with net charge of zero
• C. Separation dependent on isoelectric point
• D. Zone electrophoresis

Answer: (B)

Isoelectric focusing works by creating a continuous pH gradient in which proteins migrate in an electric field until they reach the pH that matches their isoelectric point, where their net charge is zero and they stop migrating. That stopping point at the pI is what concentrates each protein at a fixed position. Therefore, the statement about proteins migrating when they have zero net charge doesn’t fit IEF, because once the net charge is zero, there’s no driving force to move them further. The other features—having a continuous pH gradient and separations based on isoelectric point—are fundamental to how IEF operates. Zone electrophoresis describes discrete zones in some electrophoresis techniques and is not the defining approach of isoelectric focusing.