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Dual-wavelength pump-probe microscopy analysis of melanin composition.

ABSTRACT: Pump-probe microscopy is an emerging technique that provides detailed chemical information of absorbers with sub-micrometer spatial resolution. Recent work has shown that the pump-probe signals from melanin in human skin cancers correlate well with clinical concern, but it has been difficult to infer the molecular origins of these differences. Here we develop a mathematical framework to describe the pump-probe dynamics of melanin in human pigmented tissue samples, which treats the ensemble of individual chromophores that make up melanin as Gaussian absorbers with bandwidth related via Frenkel excitons. Thus, observed signals result from an interplay between the spectral bandwidths of the individual underlying chromophores and spectral proximity of the pump and probe wavelengths. The model is tested using a dual-wavelength pump-probe approach and a novel signal processing method based on gnomonic projections. Results show signals can be described by a single linear transition path with different rates of progress for different individual pump-probe wavelength pairs. Moreover, the combined dual-wavelength data shows a nonlinear transition that supports our mathematical framework and the excitonic model to describe the optical properties of melanin. The novel gnomonic projection analysis can also be an attractive generic tool for analyzing mixing paths in biomolecular and analytical chemistry.

SUBMITTER: Thompson A

PROVIDER: S-EPMC5104978 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Histotripsy produces tissue fractionation through dense energetic bubble clouds generated by short, high-pressure, ultrasound pulses. When using pulses shorter than 2 cycles, the generation of these energetic bubble clouds only depends on where the peak negative pressure (P-) exceeds the intrinsic threshold of the medium (26 to 30 MPa in soft tissue with high water content). This paper investigates a strategic method for precise lesion generation in which a low-frequency pump pulse is applied to enable a sub-threshold high-frequency probe pulse to exceed the intrinsic threshold. This pump-probe method of controlling a supra-threshold volume can be called dual-beam histotripsy. A 20-element dual-frequency (500-kHz and 3-MHz elements confocally aligned) array transducer was used to generate dual-beam histotripsy pulses in red blood cell phantoms and porcine hepatic tissue specimens. The results showed that when sub-intrinsic-threshold pump (500-kHz) and probe (3-MHz) pulses were applied together, dense bubble clouds (and resulting lesions) were only generated when their peak negative pressures combined constructively to exceed the intrinsic threshold. The smallest reproducible lesion varied with the relative amplitude between the pump and probe pulses, and, with a higher proportion of the probe pulse, smaller lesions could be generated. When the propagation direction of the probe pulse relative to the pump pulse was altered, the shape of the produced lesion changed based on the region that exceeded intrinsic threshold. Because the low-frequency pump pulse is more immune to attenuation and aberrations, and the high-frequency probe pulse can provide precision in lesion formation, this dual-beam histotripsy approach would be very useful in situations in which precise lesion formation is required through a highly attenuative and aberrative medium, such as transcranial therapy. This is particularly true if a small low-attenuation acoustic window is available for the high-frequency probe transducer.

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Dual-wavelength pump-probe microscopy analysis of melanin composition.

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