Journal: Journal of biophotonics
Changes in finger tissue blood volume (TBV) measured by light transmission and in laser Doppler flow (LDF) were obtained during long breathing (of 12 s period) and associated with the respiratory phases, inspiration and expiration. For fifteen out of sixteen subjects TBV and LDF started to decrease 0-2 s after the start of expiration and increased during inspiration but the start of increase occurred before the start of inspiration, showing that the respiratory-induced changes in TBV and LDF are mainly associated with the expiration. Decrease of TBV and LDF after expiration was also found during the inspiratory gasps (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).
Raman spectra and images of a living L929 (NCTC) cell have been measured with 532 nm excitation. Both reduced and oxidized forms of cytochromes b and c (cyt b and cyt c) have been observed in situ without any pretreatment. The redox states of cyts b and c have been assessed quantitatively with a spectral analysis. It has been found that reduced cyt c is more abundant than oxidized cyt c, while oxidized cyt b is slightly more abundant than reduced cyt b in a living cell. (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).
Tardigrades are microscopic metazoans which are able to survive extreme physical and chemical conditions by entering a stress tolerant state called cryptobiosis. At present, the molecular mechanisms behind cryptobiosis are still poorly understood. We show that surface enhanced Raman scattering supported by plasmonic gold nanoparticles can measure molecular constituents and their local distribution in live tardigrades. Surface enhanced Raman signatures allow to differentiate between two species and indicate molecular structural differences between tardigrades in water and in a dry state. This opens new avenues for exploring cryptobiosis by studying molecular changes in live cryptobiotic organisms. (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).
Low-level laser therapy (LLLT) has been extensively employed to improve epithelial wound healing, though the exact response of epithelium maturation and stratification after LLLT is unknown. Thus, this study aimed to assess the in vitro growth and differentiation of keratinocytes (KCs) and in vivo wound healing response when treated with LLLT. Human KCs (HaCaT cells) showed an enhanced proliferation with all the employed laser energy densities (3, 6 and 12 J/cm(2) , 660 nm, 100 mW), together with an increased expression of Cyclin D1. Moreover, the immunoexpression of proteins related to epithelial proliferation and maturation (p63, CK10, CK14) all indicated a faster maturation of the migrating KCs in the LLLT-treated wounds. In that way, an improved epithelial healing was promoted by LLLT with the employed parameters; this improvement was confirmed by changes in the expression of several proteins related to epithelial proliferation and maturation. Immunofluorescent expression of cytokeratin 10 (red) and Cyclin D1 (green) in (A) Control keratinocytes and (B) Low-level laser irradiated cells. Blue color illustrates the nuclei of the cells (DAPI staining).
Moderate heating of such collagenous tissues as cornea and cartilages by IR-laser irradiation is an emerging technology for non-destructive modification of the tissue shape and microstructure for a variety of applications in ophthalmology, otolaryngology, etc. Post-irradiation high-resolution microscopic examination indicates the appearance of microscopic either spheroidal or crack-like narrow pores depending on the tissue type and irradiation regime. Such examinations usually require special tissue preparation (e.g., staining, drying that affect microstructure themselves) and are mostly suitable for studying individual pores, whereas evaluation of their averaged parameters, especially in situ, is challenging. Here, we demonstrate OCT’s ability to visualize areas of pore initiation and evaluate their averaged properties by combining visualization of residual irradiation-induced tissue dilatation and evaluation of the accompanying Young-modulus reduction by OCT-based compressional elastography. We show that the averaged OCT-based data obtained in situ fairly well agree with the microscopic examination results. The results obtained develop the basis for effective and safe applications of novel non-destructive laser technologies of tissue modification in clinical practice. This article is protected by copyright. All rights reserved.
This article describes a rapid, simple and cost-effective technique that could lead to a screening method for colitis without the need for biopsies or in vivo measurements. This screening technique includes the testing of serum using Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) spectroscopy for the colitis-induced increased presence of mannose. Chronic (Interleukin 10 knockout) and acute (Dextran Sodium Sulphate-induced) models for colitis are tested using the ATR-FTIR technique. Arthritis (Collagen Antibody Induced Arthritis) and metabolic syndrome (Toll like receptor 5 knockout) models are also tested as controls. The marker identified as mannose uniquely screens and distinguishes the colitic from the non-colitic samples and the controls. The reference or the baseline spectrum could be the pooled and averaged spectra of non-colitic samples or the subject’s previous sample spectrum. This shows the potential of having individualized route maps of disease status, leading to personalized diagnosis and drug management.
The secondary structure change of the Abeta peptide to beta-sheet was proposed as an early event in Alzheimer’s disease. The transition may be used for diagnostics of this disease in an early state. We present an Attenuated Total Reflection (ATR) sensor modified with a specific antibody to extract minute amounts of Abeta peptide out of a complex fluid. Thereby, the Abeta peptide secondary structure was determined in its physiological aqueous environment by FTIR-difference-spectroscopy. The presented results open the door for label-free Alzheimer diagnostics in cerebrospinal fluid or blood. It can be extended to further neurodegenerative diseases. An immunologic ATR-FTIR sensor for Abeta peptide secondary structure analysis in complex fluids is presented.
Commercial microscopy systems make use of tandem scanning i.e. either slow or fast scanning. We constructed, for the first time, an advanced control system capable of delivering a dynamic line scanning speed ranging from 2.7 kHz to 27 kHz and achieve variable frame rates from 5 Hz to 50 Hz (512 × 512). The dynamic scanning ability is digitally controlled by a new customized open-source software named PScan1.0. This permits manipulation of scanning rates either to gain higher fluorescence signal at slow frame rate without increasing laser power or increase frame rates to capture high speed events. By adjusting imaging speed from 40 Hz to 160 Hz, we capture a range of calcium waves and transient peaks from soma and dendrite of single fluorescence neuron (CAL-520AM). Motion artifacts arising from respiratory and cardiac motion in small animal imaging reduce quality of real-time images of single cells in-vivo. An image registration algorithm, integrated with PScan1.0, was shown to perform both real time and post-processed motion correction. The improvement is verified by quantification of blood flow rates. This work describes all the steps necessary to develop a high performance and flexible polygon-mirror based multiphoton microscope system for in-vivo biological imaging.
Fluorescence lifetime measurements can provide quantitative readouts of local fluorophore environment and can be applied to biomolecular interactions via Förster resonant energy transfer (FRET). Fluorescence lifetime imaging (FLIM) can therefore provide a high content analysis (HCA) modality to map protein-protein interactions (PPIs) with applications in drug discovery, systems biology and basic research. We present here an automated multiwell plate reader able to perform rapid unsupervised optically sectioned FLIM of fixed and live biological samples and illustrate its potential to assay PPIs through application to Gag protein aggregation during the HIV life cycle. We demonstrate both hetero-FRET and homo-FRET readouts of protein aggregation and report the first quantitative evaluation of a FLIM HCA assay by generating dose response curves through addition of an inhibitor of Gag myristoylation. Z ‘ factors exceeding 0.6 are realised for this FLIM FRET assay. (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim).
Continuous monitoring of intrapulse measurement of blood flow in humans is currently not achievable with clinically available instruments. In this paper, we demonstrate a method of measuring the instantaneous variations in flow during pulsatile blood flow with an optical flow sensor comprising a fiber Bragg grating sensor and illumination from a 565 nm Light-Emitting-Diode. The LED illumination heats the blood and fluctuations in temperature, due to variations in flow, are detected by the fiber sensor. A set of experiments at different flow rates (20 to 900 mL/min) are performed in a simulated cardiac circulation setup with pulsatile flow. Data are compared with an in-line time of flight ultrasound flow sensor. Our results show that the optical and ultrasonic signals correlate with Pearson coefficients ranging from -0.83 to -0.98, dependent on the pulsatile frequency. Average flow determined by ultrasound and the optical fiber sensor showed a parabolic relationship with R2 = 0.99. An abrupt step change in flow induced by occlusion and release of the circuit tubing demonstrated that the optical fiber and ultrasound sensor had similar response. The method described is capable of intrapulse blood flow measurement under pulsatile flow conditions, with potential applications in medicine where continuous blood flow sensing is desired. This article is protected by copyright. All rights reserved.