Discovering common origins of trace evidential textile fibers can be a challenging task when fiber structure or dye composition does not provide exclusive identifying information. Introduction of new chemical species after mass production and distribution of a textile may be exploited to trace its history and identify the origin of its fibers. In this article, fluorescence microscopy is used to examine the alteration in the fluorescence spectral fingerprint of single fibers resulting from exposure to commonly used detergents that contain fluorescent whitening agents. Dyed acrylic, cotton, and nylon fibers were laundered and the spectral contribution of the detergent on single fibers was quantified and shown to reach a maximum after five sequential washes; some detergents showed statistically meaningful differences to fiber spectra after only a single wash. Principal component cluster analysis was used to determine that the spectra of laundered fibers are distinct from the spectra of dyed, unwashed cotton or nylon, but not acrylic, fibers.