Background We investigated the expression status of AGBL2 and its inhibitor latexin in breast cancer stem cells and its clinical implications in order to lay a foundation for managing breast cancer. Methods CD44+/CD24- tumor cells (CSC) from clinical specimens were sorted using flow cytometry. AGBL2 expression status was detected in CSC and 126 breast cancer specimens by western blot and immunohistochemistry staining. The relationship between the AGBL2 protein and clinicopathological parameters and prognosis was subsequently determined. Result As a result, CSC are more likely to generate new tumors in mice and cell microspheres that are deficient in non-obese diabetic/severe combined immunodeficiency mice (NOD/SCID) compared to the control group. The AGBL2 protein was expressed higher in CSC induced to epithelial to mesenchymal transition (EMT) when compared to the control cells, and was found to be related to CSC chemotherapy resistance. After Spearman regression correlation analysis, AGBL2 was observed to be related to clinical stage, histological stage, and lymph node metastasis. In the Cox regression test, the AGBL2 protein was detected as an independent prognostic factor. Through immunoprecipitation, AGBL2 and latexin could form immune complexes. Conclusions These results demonstrate that AGBL2 is a latexin- interacting protein that regulates the tubulin tyrosination cycle and is a potential target for intervention.