Label-Free Saturated Structured Excitation Microscopy

Micro- and nanoscale chemical and structural heterogeneities, whether they are intrinsic material properties like grain boundaries or intentionally encoded via nanoscale fabrication techniques, pose a challenge to current material characterization methods. To precisely interrogate the electronic structure of these complex materials systems, spectroscopic techniques with high spatial resolution are required. However, conventional optical microscopies are limited to probe volumes of similar to 200 nm due to the diffraction limit of visible light. While a variety of sub-diffraction-limited techniques have been developed, many rely on fluorescent contrast agents. Herein we describe label-free saturated structured excitation microscopy (LF-SSEM) applicable to nonlinear imaging approaches such as stimulated Raman and pump-probe microscopy. By exploiting the nonlinear sample response of saturated excitation, LF-SSEM provides theoretically limitless resolution enhancement without the need for a photoluminescent sample.