How it works
The dwell volume is everything between the point where the solvents mix and the head of the column: mixer, pump heads, tubing and, on some systems, the autosampler loop. It delays the gradient by VD/F minutes and can range from under 0.1 mL on modern UHPLC systems to more than 2 mL on older HPLCs.
When the dwell volume changes, early-eluting peaks shift most and selectivity between them can change. USP <621> allows adjusting the initial isocratic hold to compensate. A larger dwell volume cannot be removed by a hold; the fix is a delayed injection, if the autosampler supports it.