Human hair is a keratinous biological fiber with well-characterized cellular composites. For a number of decades, hair fiber has been the subject of intense scientific research in dermatology and cosmetology.1–3 In particular, structural characterization of hair has been of special interest because the morphological features of hair have provided important biophysical clues for early diagnosis of skin disease4 and breast cancer,5 cosmetic assessment,6 and forensic examination.7 Classically, ultrastructural observation of human hair has been widely performed by using scanning electron microscopy (SEM) and transmission electron microscopy (TEM).8,9 Although the microscopic assays are efficient and well-established, the sample preparation is time-consuming and laborious. It even may deform the intrinsic hair structures during cutting, vacuum evaporating, gold coating, and chemical treatments. Atomic force microscopy (AFM) and confocal laser scanning microscopy (CLSM) have emerged as alternatives for noninvasive examination of the hair structure.10–12 These techniques, compared with the electron microscopy techniques, offer three-dimensional (3-D) cellular images of the hairs with minimal sample preparation. However, AFM is only available for hair surface topography,10 and CLSM needs additional fluorescence marking of the hair layers for internal exploration.6,11,12 Further, in confocal imaging, some fluorescence signals in the regions away from the focal point might pass through the pinhole, thereby increasing emission background. Phototoxic effect of fluorophore with the laser illumination also may hamper hair cell viability. Cross-polarized CLSM (CP-CLSM), a variant of the confocal imaging regime, has provided contrast-enhanced hair imaging using polarization dependence of birefringent hair structures without aid of any chemical labeling.13,14 However, use of the technique is rather specialized in fully keratinized hair structures like cortex. Multiphoton microscopy such as two-photon excitation microscopy (TPEM) or second harmonic generation microscopy (SHM) has been employed for internal investigation of the hair.15,16 Its nonlinear excitation using a near-infrared light allowed high-resolution fluorescence imaging of the hair with long penetration depth and a small risk of photo damage. Recent effort to image the healthy state of hair without any processing or treatment has been conducted by using hard x-ray microscopy.17 The submicron spatially resolved phase contrast image showed the internal details of the hair shaft. But the long radiation of x-ray () onto the hair also might induce morphological change to the hair,18 and its projection view is not suitable to observe hair structures in depth.