Millimetre-deep micrometre-resolution vibrational imaging by shortwave infrared photothermal microscopy

Abstract

Deep tissue chemical imaging has a vital role in biological and medical applications. Current approaches suffer from water absorption and tissue scattering, which limits imaging depth to hundreds of micrometres. The shortwave infrared spectral window allows deep tissue imaging but typically features unsatisfactory spatial resolution or low detection sensitivity. Here we present a shortwave infrared photothermal (SWIP) microscope for millimetre-deep vibrational imaging with micrometre lateral resolution. By pumping the overtone transition of carbon–hydrogen bonds and probing the subsequent photothermal lens with shortwave infrared light, SWIP can obtain chemical contrast from 1 μm polymer particles located at 800 μm depth in a highly scattering phantom. The amplitude of the SWIP signal is shown to be 63 times larger than that of the optically probed photoacoustic signal. We further demonstrate that SWIP can resolve intracellular lipids across an intact tumour spheroid and the layered structure in thick liver, skin, brain and breast tissues. SWIP microscopy fills a gap in vibrational imaging with subcellular resolution and millimetre-level penetration, which heralds broad potential for life science and clinical applications. Shortwave infrared photothermal microscopy enables chemical imaging at millimetre depths with a micrometre spatial resolution in tissue-mimicking phantoms, intact tumour spheroids and various biological tissues.