In vivo single human sweat gland activity monitoring using coherent anti-Stokes Raman scattering and two-photon excited autofluorescence microscopy.

BACKGROUND: Eccrine sweat secretion is of central importance for control of body temperature. Although the incidence of sweat gland dysfunction might appear of minor importance, it can be a real concern for people with either hypohidrosis or hyperhidrosis. However, sweat gland function remains relatively poorly explored. OBJECTIVES: To investigate the function of single human sweat glands. METHODS: We describe a new approach for noninvasive imaging of single sweat gland activity in human palms in vivo up to a depth of 100 µm, based on nonlinear two-photon excited autofluorescence (TPEF) and coherent anti-Stokes Raman scattering (CARS). RESULTS: These techniques appear to be useful compared with approaches already described for imaging single sweat gland activity, as they allow better three-dimensional spatial resolution of sweat pore inner morphology and real-time monitoring of individual sweat events. By filling the sweat pore with oil and tuning the CARS contrast at 2845 cm(-1) , we imaged the ejection of sweat droplets from a single sweat gland when oil is pushed out by sweat flow. On average, sweat events lasted for about 30 s every 3 min under the conditions studied. On the other hand, about 20% of sweat glands were found inactive. TPEF and CARS were also used to study, at the single pore level, the antiperspirant action of aluminium chlorohydrate (ACH) and to reveal, for the first time in vivo, the formation of a plug at the pore entrance, in agreement with reported ACH antiperspirant mechanisms. CONCLUSIONS: Although data were acquired on human palms, these techniques show great promise for a better understanding of sweat secretion physiology and should be helpful to improve the efficacy of antiperspirant formulations.

Terahertz imaging applied to cancer diagnosis.

We report on terahertz (THz) time-domain spectroscopy imaging of 10 microm thick histological sections. The sections are prepared according to standard pathological procedures and deposited on a quartz window for measurements in reflection geometry. Simultaneous acquisition of visible images enables registration of THz images and thus the use of digital pathology tools to investigate the links between the underlying cellular structure and specific THz information. An analytic model taking into account the polarization of the THz beam, its incidence angle, the beam shift between the reference and sample pulses as well as multiple reflections within the sample is employed to determine the frequency-dependent complex refractive index. Spectral images are produced through segmentation of the extracted refractive index data using clustering methods. Comparisons of visible and THz images demonstrate spectral differences not only between tumor and healthy tissues but also within tumors. Further visualization using principal component analysis suggests different mechanisms as to the origin of image contrast.

Investigation of dyed human hair fibres using apertureless near-field scanning optical microscopy.

We present the first studies of dyed human hair fibres performed with an apertureless scanning near-field optical microscope. Samples consisted of 5-microm-thick cross-sections, the hair fibres being bleached and then dyed before being cut. Hair dyed with two molecular probes diffusing deep inside the fibre or mainly spreading at its periphery were investigated at a wavelength of 655 nm. An optical resolution of about 50 nm was achieved, well below the diffraction limit; the images exhibited different optical contrasts in the cuticle region, depending on the nature of the dye. Our results suggest that the dye that remains confined at the hair periphery is mainly located at its surface and in the endocuticle.

Analysis of the measured signals in apertureless near-field optical microscopy.

We present an analytical model able to explain the optical signal recorded during our experimental approach curves in the infrared at a wavelength lambda=10.6 microm, with a home-made apertureless near-field scanning optical microscope ANSOM. This model uses classical electrodynamics to calculate the scattering cross section of the oscillating tip, considered as a dipole, and its dielectric image in the sample as a function of the tip-sample separation from the near-field to the far-field regime. The dipoles are placed in a non-uniform electric field because of the standing wave arising from the interference between the incident and the specular laser beams. We also added a background field coming from a scatterer on the surface in order to account for zeroing of the optical signal for particular tip-sample separation and interference patterns.

Apertureless near-field scanning optical microscope working with or without laser source.

An apertureless near-field scanning optical microscope (ANSOM), used indifferent configurations, is presented. Our versatile home-made setup, based on a sharp tungsten tip glued onto a quartz tuning fork and working in tapping mode, allows to perform imaging over a broad spectral range. We have recorded optical images in the visible (wavelength, lambda = 655 nm) and in the infrared (lambda = 10.6 microm), proving that the setup routinely achieves an optical resolution of <50 nm regardless of the illumination wavelength. We have also shown optical images recorded in the visible (lambda = 655 nm) in an inverted configuration where the tip does not perturb the focused spot of the illumination laser. Approach curves as well as image profiles have revealed that on demodulating the optical signal at higher harmonics, we can obtain an effective probe sharpening which results in an improvement of the resolution. Finally, we have presented optical images recorded in the infrared without any illumination, that is, the usual laser source is replaced by a simple heating of the sample. This has shown that the ANSOM can be used as a near-field thermal optical microscope (NTOM) to probe the near field generated by the thermal emission of the sample.

Apertureless scanning near-field fluorescence microscopy in liquids.

We show that apertureless scanning near-field optical microscopes that use sharp vibrating conical tips can be operated in liquid environments. We have investigated the damping of the tip oscillation as a function of its shape and as a function of its depth under the liquid surface. The degradation of the quality factor from 150 in air down to 15 in liquid does not impede to perform topographic and optical measurements with a very good sensitivity. As an example of application, we present near-field fluorescence images of dye-doped polystyrene spheres immersed in a liquid.