Step 1: State the concept.
Drug protein binding measures how much of a drug attaches to plasma proteins like albumin. To measure it we need a method that separates free drug from protein-bound drug.
Step 2: Recall the standard technique.
Equilibrium dialysis is the classic method. A semi-permeable membrane separates the protein-drug solution from a buffer; only free drug crosses while protein and bound drug stay behind.
Step 3: Reason through the measurement.
Once equilibrium is reached, measuring free drug concentration on the buffer side lets us calculate the bound fraction. This direct separation is why dialysis is the method of choice.
Step 4: Eliminate chromatography.
Option 1 (chromatography) is mainly an analytical separation and quantification tool; it is not the standard, defining method for binding studies, so it is rejected.
Step 5: Eliminate phase separation and Langmuir's method.
Option 3 (phase separation) is not a recognised protein binding assay, and option 4 (Langmuir's method) relates to surface adsorption isotherms, not plasma protein binding, so both are discarded.
Step 6: Conclude.
Drug protein binding is determined by dialysis (equilibrium dialysis), which is option 2.
\[ \boxed{\text{Option 2: Dialysis}} \]