ZnT2 imports zinc (Zn) into vesicles for secretion from mammary epithelial cells (MEC) into milk. Four mutations in ZnT2 have been identified that abrogate Zn secretion into milk, resulting in severe neonatal Zn deficiency; however, the role of genetic variation in ZnT2 on MEC function and milk Zn level has not been explored. DNA from buccal swabs and milk from healthy, exclusively breastfeeding women (n=54) were obtained at 4 mo postpartum. Milk Zn level was stratified into tertiles (high, >1.7 μg/mL; normal, 1‐1.6 μg/mL; low, <0.9 μg/mL) and the ZnT2 coding region was sequenced. ~50% of women had genetic variation in ZnT2. Four SNPs (rs142587047, rs149340896, rs150251854, rs145406127) and 17 novel non‐synonymous variants were identified and only found in women in the high and low tertiles. 8 select variants were generated to functionally characterize defects in MECs in vitro. Variants responsible for low Zn secretion were confined to the ER/Golgi apparatus and mitochondria and induced apoptosis and G1 arrest. Variants responsible for high Zn secretion were mislocalized to the cell membrane and induced G1 arrest and entry into S and G2 phase. Altered function corresponded to a redistribution of subcellular Zn as assessed by FluoZin3. This suggests that genetic variation in ZnT2 is common and affects ZnT2 function and milk Zn levels. Defects in ZnT2 may compromise infant health and modify a woman’s risk for breast disease.