Widefield super-resolution photothermal infrared spectroscopy and imaging of autofluorescent biomaterials and photosynthetic microorganisms

A variety of biological materials and photosynthetic organisms exhibit strong autofluorescence emission under ultraviolet excitation and the autofluorescent emission has a very strong temperature dependence, of order 1%/K. Taking advantage of this property, we have demonstrated high-speed, super-resolution photothermal infrared spectroscopy and chemical imaging of autofluorescent biomaterials and organisms using camera-based widefield autofluorescence detection. Illuminating a sample with pulses of infrared light from a tunable laser source causes periodic localized sample temperature increases that result in a significant decrease in autofluorescent emission. A conventional s-CMOS camera is used to detect localized variations in autofluorescent emission over a wide area as an indicator of localized IR absorption. IR absorption image stacks can be acquired over a range of infrared wavelengths, including the fingerprint range, enabling extraction of localized IR absorption spectra. We have applied widefield FE-PTIR to analysis of autofluorescent biological materials including collagen, leaf tissue, and photosynthetic organisms including diatoms and green microalgae cells. We have also demonstrated chemical imaging of live microalgae in water.