Lock-in infrared thermography was used to estimate the fatigue limit for Ti6Al4V alloy at room temperature. The method detected infrared emitted from specimen during cyclic loading, i.e., temperature change related to frequency ( f ) of the loading. The temperature change contained reversible component and irreversible component, which related to thermos-elastic effect and plastic deformation, respectively. The latter component was divided from the former one by lock-in analysis to estimate fatigue limit. Estimated fatigue limits corresponded to those obtained from conventional fatigue tests at several stress ratios. A new assessment line was identified as, where • a represents the stress amplitude, • w signifies the fatigue limit, • m denotes mean stress, • B expresses tensile strength, and the n exponent for the alloy is about 2.3. The non-failure region of the new diagram was smaller than that of the modified Goodman diagram because of a reduced fatigue limit at near-zero stress ratios in the titanium alloy.