Nanosecond-resolution photothermal dynamic imaging via MHZ digitization and match filtering

Abstract

Photothermal microscopy has enabled highly sensitive label-free imaging of absorbers, from metallic nanoparticles to chemical bonds. Photothermal signals are conventionally detected via modulation of excitation beam and demodulation of probe beam using lock-in amplifier. While convenient, the wealth of thermal dynamics is not revealed. Here, we present a lock-in free, mid-infrared photothermal dynamic imaging (PDI) system by MHz digitization and match filtering at harmonics of modulation frequency. Thermal-dynamic information is acquired at nanosecond resolution within single pulse excitation. Our method not only increases the imaging speed by two orders of magnitude but also obtains four-fold enhancement of signal-to-noise ratio over lock-in counterpart, enabling high-throughput metabolism analysis at single-cell level. Moreover, by harnessing the thermal decay difference between water and biomolecules, water background is effectively separated in mid-infrared PDI of living cells. This ability to nondestructively probe chemically specific photothermal dynamics offers a valuable tool to characterize biological and material specimens.

Ultrafast Widefield Mid-Infrared Photothermal Heterodyne Imaging

“…The inclusion of the higher harmonics in the demodulation improved the SNR by a factor of ∼2, since all MIP signal components were recovered (Figure ). This was also demonstrated in a previous work using point-scan imaging …”

“…The ability to record decay profiles is an important feature that has proven useful for instrument design and optimization, theory development, signal simulation, and target compound characterization. 15,26 Here, it is demonstrated for K-FeCy microparticles that the thermal decay time decreases with particle size, most likely because the effective transfer surface area between the specimen and the surrounding increases. In general, K-FeCy exhibits longer thermal decay times than polymer beads of the same size, 26 even when suspended in 1-propanol, which facilitates heat dissipation.…”

“…This was also demonstrated in a previous work using point-scan imaging. 15 Based on the Abbe resolution equation R x, y = λ/2NA, the theoretical diffraction limit of the current WIPH system is 411 nm. As confirmed with the help of the test target, the spatial coverage of one camera pixel, which is defined by the objective magnification (40×), the focal length of the tube lens (200 mm), and the pixel size (20 μm), is ∼500 nm.…”

“…Since a LIA usually demodulates only one of the (lower) harmonics, the MIP signal amplitude is not maximized, and the nonlinear photothermal decay cannot be accurately resolved. This led to the development of multiharmonic demodulation MIP schemes, such as photothermal dynamics imaging (PDI) . PDI makes use of high bandwidth detection to obtain nanosecond temporal resolution per pixel within a single pulse excitation and enables retrieval of the thermal decay profile, which can improve the chemical contrast in complex samples and provide valuable physical properties of the target.…”

Bond-selective intensity diffraction tomography

“…The emerging mid-infrared photothermal (MIP) microscopy 18 inherits IR absorption spectroscopy's advantages but circumvents conventional IR micro-spectroscopy's low-resolution and slow speed limit. MIP microscopy provides diffraction-limited resolution at the visible band using a visible probe beam and is compatible with both point-scanning [25][26][27][28] and wide-field [29][30][31][32][33][34] configuration. Yet, existing MIP microscopy suffer from slow volumetric imaging speed and low depth resolution.…”