Question

PART A: 137 nmol of an antibody that binds strongly to the antigen protein G is covalently bound to the support of an affinity column with a void volume of 1.95 mL. The association equilibrium constant (K_A) for the binding of protein G to the antibody is 5.19 × 10⁸ M^-1 at pH 7.2. Calculate the retention factor for protein G on this column at pH 7.2. k=?

PART B: At pH 7.2 and a flow rate of 1.00 mL/min, how many minutes would it take protein G to elute from this column? t_r=? min

PART C: If a pH 3.5 elution buffer is applied to the column, protein G elutes at 3.12 min using a flow rate of 1.00 mL/min. Under these conditions, what is the association equilibrium constant between protein G and the antibody? K_A=? M^-1

Answer 1