During the identification of single fiber evidence, color evaluation is one of the most important examinations. Colors are usually first evaluated by ultraviolet-visible-microspectrophotometry (UV-vis-MSP). If possible, thin-layer chromatography (TLC) or high performance liquid chromatography (HPLC) are conducted after the extraction of dyestuffs from the single fiber. In this report, three woolen single fibers were dyed by similarly colored dyes with structurally different yellow acid dyes and were used to evaluate the discriminatory abilities of UV-vis-MSP and TLC.