Multimodal compressive microscopy of fluorescence dynamics and transmittance spectra

Abstract

Absorption spectroscopy and photoluminescence (PL) decay measurement count among the most used spectroscopic methods to characterize optical materials. Therefore, an extension of these techniques into the imaging mode – hyperspectral imaging (HSI) and fluorescence lifetime imaging (FLIM) – is immensely useful and commonly used. The measurement of transmittance HSI and FLIM is seldom combined in a single system due to the necessity to use very different working conditions. For HSI, high light intensity needs to illuminate a sample as its image is typically scanned across a slit. On the contrary, the most common FLIM technique, time-correlated single-photon counting (TCSPC), uses low-light conditions and laser beam scanning. We present a versatile multimodal microscopic system that combined transmittance HSI with FLIM using a single setup based on compressed sensing, namely on the so-called single-pixel camera (SPC). This method makes it possible to reconstruct an image by using only a fraction of the measurements necessary compared to the number of pixels. The use of the SPC concept allowed us to create multidimensional HS and FLIM maps, which are inherently matched. Moreover, their spectral and temporal resolution follows the resolution of the used spectrometer and TCSPC setup. Our system is also highly versatile with respect to image magnification and field of view. Owing to the use of a digital micromirror device, we can switch without any modification of the setup, from the large field of view (3.5 × 2.6 mm) to the imaging on the micrometer scale limited only by the microscope lens. We demonstrate the functionality of the setup in both HS and FLIM mode on the testing samples of fluorescent dyes, luminophore (LuAG:Ce), and the cells of Convallaria.