Background To investigate the differential expression levels of thymosin β10 (Tβ1O) and the corresponding changes of actin filament organization in human tumor cell lines with different metastatic potential.Methods Four groups of nine human tumor cell lines with different metastatic potential were analyzed for the amount of Tβ10 mRNAs by Northern blot and for their peptide expression levels by immunohistochemistry. The filamentous actin (F-actin)was observed by staining of TRITC-phalloidin to detect changes in actin organization. Results In comparison with non-/weakly metastatic counterparts, TβIO was upregulated in highly metastatic human lung cancer, malignant melanoma and breast cancer cell lines. Staining of TRITC-phalloidin revealed less actin bundles, a fuzzy network of shorter filaments and some F-actin aggregates in the highly metastatic tumor cells. Meanwhile, the actin filaments were robust and orderly arranged in the non-/weakly metastatic cancer cell lines.Conclusion Tβ10 levels correlate positively with the metastatic capacity in human tumors currently examined. The increasing metastatic potential of tumor cells is accompanied by a loss of F-actin,poorly arranged actin skeleton organizations and presence of F-actin aggregates. There is a consistent correlation between the elevated TβIO expression and the disrupted actin skeleton.