Integrated photothermal flow cytometry in vivo.
ABSTRACT The capability of integrated flow cytometry to detect, in real time, moving cells in their natural states in vivo is demonstrated in a study of circulating red and white blood cells in lymph and blood flow of rat mesentery. This system combines dual pump-probe photothermal (PT) techniques, such as PT imaging, the PT thermolens method, and PT velocimetry, with high-resolution (up to 0.3 microm), high-speed (up to 1000 fps) transmission digital microscopy (TDM) and fluorescence imaging. All PT techniques are based on irradiation of cells in rat mesenteric microvessels with a spectrally tunable laser pulse (420 to 570 nm, 8 ns, 0.1 to 300 microJ) and on detection of temperature-dependent variations of the refractive index with a second continuous probe laser beam (633 nm, 1.4 mW). We focus on intravital monitoring of the integral PT response from single, moving, unlabeled cells (from 100 to 500 cells in one measurement). Potential in vivo applications of this new optical tool, called PT flow cytometry (PTFC), are discussed, including identification of selected cells with differences in natural absorptive properties and sizes, determination of laser-induced cell damage, estimation of flow velocity, and monitoring of circulating cells labeled with PT probes.
SourceAvailable from: Dmitry A Nedosekin[Show abstract] [Hide abstract]
ABSTRACT: The main examples of the application of thermal lens spectrometry and thermal lens microscopy as highly sensitive power-based thermooptical (photothermal) methods of molecular absorption spectroscopy to photometric, chromatographic, and electromigration methods of analysis and to solving problems related to the use of microfluid chips are considered. The fundamentals and characteristics of the thermooptical methods and instruments and the problems and prospects of their development are discussed. Examples are provided for illustrating the sensitivity and selectivity of combined methods and multidisciplinary studies performed with the aid of thermal lens spectrometry.Journal of Analytical Chemistry 03/2015; 70(3):249-276. DOI:10.1134/S1061934815030168 · 0.81 Impact Factor
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ABSTRACT: A thin photothermal (PT) endoscope (similar to 80 mu m) for the noninvasive/minimally invasive hybrid-optical diagnosis of biological specimens is demonstrated. The technique has the unique advantage that the pump laser delivery fiber itself acts as the thermal wave sensor, which is a Bragg grating. It detects only the conductive component of the PT signal, thus enabling an emissivity independent measurement. The device is slidable through a syringe needle and PT analysis of exposed organs with limited accessibility for conventional PT techniques, and constricted regions can be examined noninvasively. For regions buried in thick tissues, a minimally invasive injection mode may be considered. Temperature measurement sensitivity is about 0.03 degrees C. The amplitude and phase channels are sensitive up to about 3 and 10 kHz, respectively. The endoscope has been used for the simultaneous estimation of flow velocity, absorption coefficient, and diffusivity for a phantom-blood flow. The endoscopically estimated values are in agreement with true flow velocities over a range of 1 to 1000 cm(-1). The endoscope has been used for the optical biopsy of goat bone marrow. (C) 2013 Society of Photo-Optical Instrumentation Engineers (SPIE)Journal of Biomedical Optics 09/2013; 18(9):7008-. DOI:10.1117/1.JBO.18.9.097008 · 2.75 Impact Factor
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ABSTRACT: Despite progress in detecting circulating tumor cells (CTCs), existing assays still have low sensitivity (1-10 CTC/mL) due to the small volume of blood samples (5-10 mL). Consequently, they can miss up to 103-104 CTCs, resulting in the development of barely treatable metastasis. Here we analyze a new concept of in vivo CTC detection with enhanced sensitivity (up to 102-103 times) by the examination of the entire blood volume in vivo (5 L in adults). We focus on in vivo photoacoustic (PA) flow cytometry (PAFC) of CTCs using label-free or targeted detection, photoswitchable nanoparticles with ultrasharp PA resonances, magnetic trapping with fiber-magnetic-PA probes, optical clearance, real-time spectral identification, nonlinear signal amplification, and the integration with PAFC in vitro. We demonstrate PAFC's capability to detect rare leukemia, squamous carcinoma, melanoma, and bulk and stem breast CTCs and its clusters in preclinical animal models in blood, lymph, bone, and cerebrospinal fluid, as well as the release of CTCs from primary tumors triggered by palpation, biopsy or surgery, increasing the risk of metastasis. CTC lifetime as a balance between intravasation and extravasation rates was in the range of 0.5-4 h depending on a CTC metastatic potential. We introduced theranostics of CTCs as an integration of nanobubble-enhanced PA diagnosis, photothermal therapy, and feedback through CTC counting. In vivo data were verified with in vitro PAFC demonstrating a higher sensitivity (1 CTC/40 mL) and throughput (up to 10 mL/min) than conventional assays. Further developments include detection of circulating cancer-associated microparticles, and super-rsesolution PAFC beyond the diffraction and spectral limits.12/2013; 5(4):1691-1738. DOI:10.3390/cancers5041691