The rise in antibiotic resistance has stimulated research into adjuvants that can improve the efficacy of broad-spectrum antibiotics. Lactoferrin is a candidate adjuvant; it is a multifunctional iron-binding protein with antimicrobial properties. It is known to show dose-dependent antimicrobial activity against Staphylococcus aureus through iron sequestration and repression of β-lactamase expression. However, S. aureus can extract iron from lactoferrin through siderophores for their growth, which confounds the resolution of lactoferrin's method of action. We measured the minimum inhibitory concentration (MIC) for a range of lactoferrin/ β-lactam antibiotic dose combinations and observed that at low doses (< 0.39 μM), lactoferrin contributes to increased S. aureus growth, but at higher doses (> 6.25 μM), iron-depleted native lactoferrin reduced bacterial growth and reduced the MIC of the β-lactam-antibiotic cefazolin. This differential behaviour points to a bacterial population response to the lactoferrin/ β-lactam dose combination. Here, with the aid of a mathematical model, we show that lactoferrin stratifies the bacterial population, and the resulting population heterogeneity is at the basis of the dose dependent response seen. Further, lactoferrin disables a sub-population from β-lactam-induced production of β-lactamase, which when sufficiently large reduces the population's ability to recover after being treated by an antibiotic. Our analysis shows that an optimal dose of lactoferrin acts as a suitable adjuvant to eliminate S. aureus colonies using β-lactams, but sub-inhibitory doses of lactoferrin reduces the efficacy of β-lactams.