Anti-centromere antibodies (ACA) can be detected by indirect immunofluorescence (IFI) or immunodot (ID). Our aim was to assess IFI/ID concordance and the clinical relevance of ACA.Patients presenting with ACA positivity by IFI on Hep-2 cells and/or by ID were included. ACAs were detected in 63 cases (2.1% of IFI+ID tests). ACAs were positive by IFI and ID in 18 cases. Discordances involved 2 cases of ACA-IFI+/ACA-ID- and 43 cases of ACA-IFI-/ACA-ID+. For these 43 cases, the nuclear fluorescence appearance (IFI) was speckled (34), nucleolar speckled (6) or homogeneous (3); ACA (ID) was weakly positive (26), positive (7) or strongly positive (10).The diagnosis of connective tissue disease (systemic scleroderma, systemic lupus erythematosus, rheumatoid arthritis) was retained in 3 patients out of 18 with clinical information. IFI on Hep-2 cells has certain limitations (operator-dependent; one fluorescence appearance may be masked by another). ID is useful for confirming antigenic specificity, within the limits of the antigens used. Interpretation of these tests depends on the clinical context.