Hyperspectral Three-Dimensional Refractive-Index Imaging Using Snapshot Optical Tomography

The refractive index serves as an intrinsic contrast in light-scattering measurements for the estimation of the size, shape, and heterogeneities of microscopic particles. The refractive index of cells, intracellu-lar organelles, and extracellular materials is an important input to the analysis of diffuse optical imaging of biological tissues. To improve the accuracy, the measurement is often performed at multiple discrete wavelengths or sometimes over a continuous wavelength range. Knowledge of the refractive index as a function of the wavelength is important, as the analysis of the acquired signal heavily relies on the relationship. Digital holographic tomography allows us to directly measure the three-dimensional (3D) refractive-index map of a heterogeneous microscopic specimen. Typically, a multitude of images are recorded for varying angles of illumination and a tomographic reconstruction algorithm is applied for the 3D reconstruction. Hyperspectral 3D refractive-index imaging has been demonstrated by combining the beam-rotation tomography with a wavelength-scanning light source; however, the data acquisition is slow due to the requirement of scanning both the orientation and the wavelength of the illumination. Recently, several strategies have been proposed to acquire the 3D tomogram in a single snapshot. Here, we combine snapshot holographic optical tomography with a wavelength-scanning laser to demonstrate hyperspectral 3D refractive-index imaging at high throughput. Using the developed system, we measure the refractive-index dispersion of polystyrene beads and single living HeLa cells.