The objective of the study was to determine the diagnostic yield of the indirect immunofluorescence (IFI) test for the detection of antibodies against HTLV-1. A diagnostic test evaluation study was performed. HTLV-1-infected MT2 cells and HTLV-1-uninfected K-562 cells were cultured; then these cells were impregnated and fixed in sheets for immunofluorescence and faced to Peruvian sera. A total of 155 sera (80 HTLV-1-positive sera and 75 sera positive for other diseases) from the Peruvian Instituto Nacional de Salud were used. In addition, the parameters of repeatability (intra-laboratory) and reproducibility (in laboratories of the Peruvian coast, mountains and jungle) of the test were evaluated. The IFI test detected the presence of antibodies against HTLV-1 reaching a sensitivity of 98.75% (95% CI: 95.69 – 100.00%), a specificity of 98.67% (95% CI: 95.40 – 100.00%) and the Kappa index was 0.975. There were no false positives or false negatives; however, one undetermined result and one non-specific result were obtained. The test showed 100% qualitative agreement when performing the repeatability and reproducibility. The results obtained are comparable to the reference test. Therefore, the IFI test had a good diagnostic performance and would be useful for the confirmation of HTLV-1.