We propose that the slightly acidic mobile phase pH at the end of the gradient results in a conforma9onal change, thus altering the surface charge distribu9on of the par9cles and in consequence their pI. VP 1 was previously reported to undergo structural changes if subjected to heat stress or low pH . The N-terminal phospholipase A2 domain of the capsid protein was shown to undergo a reversible unfolding/refolding process at a pH of 5.5 -4.0 which leaves the capsid intact.