Abstract
.Significance: Mid-infrared (IR) imaging based on the vibrational transition of biomolecules provides good chemical-specific contrast in label-free imaging of biology tissues, making it a popular tool in both biomedical studies and clinical applications. However, the current technology typically requires thin and dried or extremely flat samples, whose complicated processing limits this technology’s broader translation.Aim: To address this issue, we report mid-IR photoacoustic microscopy (PAM), which can readily work with fresh and thick tissue samples, even when they have rough surfaces.Approach: We developed a transmission-mode mid-IR PAM system employing an optical parametric oscillation laser operating in the wavelength range from 2.5 to . Due to its high sensitivity to optical absorption and the low ultrasonic attenuation of tissue, our PAM achieved greater probing depth than Fourier transform IR spectroscopy, thus enabling imaging fresh and thick tissue samples with rough surfaces.Results: In our spectroscopy study, the symmetric stretching at (3508 nm) was found to be an excellent source of endogenous contrast for lipids. At this wavenumber, we demonstrated label-free imaging of the lipid composition in fresh, manually cut, and unprocessed tissue sections of up to 3-mm thickness.Conclusions: Our technology requires no time-consuming sample preparation procedure and has great potential in both fast clinical histological analysis and fundamental biological studies.
Highlights
Imaging technologies exploiting the vibrational transition of biomolecules,[1,2] such as Fourier transform infrared (FTIR) spectroscopy[3] and stimulated Raman scattering (SRS) imaging,[4] provide rich and specific information about tissue’s biochemical composition in a label-free manner.[5,6,7,8,9,10]
He et al.: Label-free imaging of lipid-rich biological tissues by mid-infrared photoacoustic microscopy it detects is heavily attenuated by biological tissue, which is highly absorbing in the mid-IR range.[9]
The output laser beam was split into two parts by a germanium (Ge) beam splitter, with one part going to the imaging arm and the other to the reference arm
Summary
Imaging technologies exploiting the vibrational transition of biomolecules,[1,2] such as Fourier transform infrared (FTIR) spectroscopy[3] and stimulated Raman scattering (SRS) imaging,[4] provide rich and specific information about tissue’s biochemical composition in a label-free manner.[5,6,7,8,9,10] Because of their ability to classify biomolecules (e.g., glycogen, proteins, lipids, or nucleic acids),[11,12] these technologies have many biomedical applications, such as analyzing clinical biopsy samples ex vivo[13,14,15,16] and studying disease progression with tissue sections taken from animal models.[17,18] their broader translation is still inhibited by certain intrinsic limitations. He et al.: Label-free imaging of lipid-rich biological tissues by mid-infrared photoacoustic microscopy it detects is heavily attenuated by biological tissue, which is highly absorbing in the mid-IR range.[9] An emerging FTIR technique, attenuated total reflection (ATR), can image thick samples and live cells in an aqueous environment.[10,22,23,24,25] it still requires time-consuming sample preparation and an expensive slicing instrument to produce an extremely flat surface,[26,27,28] because its maximum probing depth is only ∼3 μm.[22,25,29] SRS imaging is capable of imaging unprocessed tissues, but it requires a complicated detection device and suffers from poor sensitivity and possible photodamage[30] since it relies on the weak Raman scattering effect.[2]
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