Linearity, interference evaluations of the performance of clinical chemistry systems, mathematical model selection for nonlinear calibration, and other assessments often involve several human sample pools with equally spaced analyte concentrations. Sequential mixing of equal volumes, first of the low and high pools to produce the middle pool, then of the low and middle pools to produce the mid-low pool, and of the high and middle pools to produce the mid-high pool, is recommended in the NCCLS EP7-P guideline for interference studies. Proportional mixing of the low and high pools to produce all of the required pool concentrations is recommended in the NCCLS EP6-P guideline for linearity studies. Mathematical analysis and computer simulation show that the sequential mixing is much more accurate and precise than the proportional mixing. Therefore, we recommend sequential mixing for clinical chemistry application.