High-speed scanless entire bandwidth mid-infrared chemical imaging

Abstract

Mid-infrared spectroscopy probes molecular vibrations to identify chemical species and functional groups. Therefore, mid-infrared hyperspectral imaging is one of the most powerful and promising candidates for chemical imaging using optical methods. Yet high-speed and entire bandwidth mid-infrared hyperspectral imaging has not been realized. Here we report a mid-infrared hyperspectral chemical imaging technique that uses chirped pulse upconversion of sub-cycle pulses at the image plane. This technique offers a lateral resolution of 15 µm, and the field of view is adjustable between 800 µm × 600 µm to 12 mm × 9 mm. The hyperspectral imaging produces a 640 × 480 pixel image in 8 s, which covers a spectral range of 640-3015 cm^-1, comprising 1069 wavelength points and offering a wavenumber resolution of 2.6-3.7 cm^-1. For discrete frequency mid-infrared imaging, the measurement speed reaches a frame rate of 5 kHz, the repetition rate of the laser. As a demonstration, we effectively identified and mapped different components in a microfluidic device, plant cell, and mouse embryo section. The great capacity and latent force of this technique in chemical imaging promise to be applied to many fields such as chemical analysis, biology, and medicine.

Fig. 1. Concept and setup of the high-speed scanless mid-infrared hyperspectral chemical imaging.

a Conceptual illustration of high-speed scanless mid-infrared (MIR) hyperspectral chemical imaging. b Mid-infrared hyperspectral chemical imaging setup. M1, M2, M3, and M4: mirrors (high reflection for ω_F), BS: beam splitter with 70% transmittance and 30% reflectance. BBO: β-BaB₂O₄ (type-I, cut angle 29˚, thickness 100 µm). CM1: dielectric concave mirror (high reflection for ω_F and ω_SHG, radius of curvature: 1.5 m), CM2: aluminium-coated concave mirror (radius of curvature: 0.5 m) with a hole of 7 mm diameter, CM3, CM4, and CM5: gold-coated concave mirrors (radius of curvature, CM3: 0.5 m, CM4: 0.4 m and CM5: 0.3 m), CR: corner reflector with two mirrors (high reflection for ω_F), LPF: long pass filter (Semrock FF01-776/LP-25, cut-on wavelength: 783 nm), SPF: short pass filter (Semrock FF01-770/SP-25, cut-on wavelength: 752 nm), Lens: achromatic lens (focal length 200 mm), M5 and M6: aluminium-coated mirrors, BPF: tunerable bandpass filter set (Semrock TBP01-704/13-25x36 and TBP01-790/12-25x36), HSI: hyperspectral imaging unit (EBA JAPAN, SIS-H-0.45nm, number of effective pixels: 640 × 480), II-HSC: a high-speed gated image intensifier unit (Hamamatsu, 10880-13F) and a high-speed camera (Phantom, VEO 410L).

Fig. 2. MIR spectrum and spatial resolution of the imaging system.

Upconverted MIR spectrum by a 100 µm and b 4.4 µm thick GaSe crystal. c Calculation results of the configuration of GaSe crystal-type. d Spectrally resolved cross-correlation between the MIR pulse and the chirped pulse. e Upconversion MIR spectra of CO₂ gas before and after retrieval. Infrared transmittance spectrum of CO₂ gas by a commercially available FT-IR. f Image of a copper mesh grid. g Intensity profile marked by the red dashed line in f. The spatial resolution is estimated as 15 μm and is defined by the edge response of 10–90% distance of the line profile. h MIR hyperspectral stability at an exposure time of 8 s. CP: chirped pulse, MIR: mid-infrared, SFG: sum frequency generation, o: ordinary ray, e: extraordinary ray, FID: free induction decay.

Fig. 3. Hyperspectral image for qualitative analysis.

a Reflected electron image of a 5 microchannels device. b Optical microscopic image of microchannels device injected with 5 samples obtained before the measurement of mid-infrared hyperspectral imaging. c Mapped the mid-infrared hyperspectral image, from left to right are glycerin, glucose, albumin, 1,2-dioleoyl-sn-glycero-3-phosphocholineglucose and soybean oil. The mapped color is a pseudo-color. d Each MIR transmittance spectrum of each sample shown in c. The black lines are infrared transmittance spectra of the 5 samples measured in a vacuum (4 Pa) by FT-IR (JASCO, FT/IR-6100). e Images corresponding to fifteen different individual wavelength components. A movie (Supplementary Movie 1) of the 2D image while the wavelength is continuously scanned over the entire mid-infrared bandwidth.

Fig. 4. Discrete frequency MIR images of water.

Water was filled into three microchannels with a wide of 200 µm and deep of 25 µm. In the middle microchannel, water starts to evaporate from the upper and the water flows due to surface tension. The water in the left and right microchannels has not yet started to evaporate, thus the MIR has been absorbed all the time and shows a dark distribution. The MIR is no longer absorbed by the water at the position after the water has flowed away, thus showing a brighter distribution. The time resolution is 0.2 ms and the exposure time is 1 µs per frame. a 6 frames shown here are images extracted every 350 frames, that is, every 70 ms. The size of the images is 1200 µm × 900 µm (640 × 480 pixels). The bit depth of the high-speed imaging of discrete wavelengths is 8 bit. b Temporal evolution of the intensity at the two marked 30 × 30 pixels. The recorded video is given in Supplementary Movie 2.

Fig. 5. MIR hyperspectral imaging and mapping of the onion (A. cepa) bulb leaves epidermal cells.

a Microscopy image illuminated by visible light. Mapped hyperspectral images of b cell wall, c cell membrane, d cell nucleus, and e cytoplasm and f their overlap were obtained by spectral analysis. The mapped color is a pseudo-color. g Infrared transmittance spectra of the cell wall, cell membrane, cell nucleus, and cytoplasm by extracted hyperspectral image. h Stained visible light microscopy image with acetocarmine solution which took after the hyperspectral image measurement. The scale bar applies to all images in Fig. 5.

Fig. 6. Hyperspectral image of a sagittal plane section of 13.5 days age mouse embryo.

a Photograph by white light illumination. Labels assigned based on anatomy and morphology: (1) diencephalon and (2) muscle. b, Mapped the mid-infrared hyperspectral image of 2 classified components. The mapped color is a pseudo-color. c, MIR transmittance spectra of 2 classes which are extracted from hyperspectral image.