Xuantao Su

Measurements of light scattering in an integrated microfluidic waveguide cytometer.

An integrated microfluidic planar optical waveguide system for measuring light scattered from a single scatterer is described. This system is used to obtain 2D side-scatter patterns from single polystyrene microbeads in a fluidic flow. Vertical fringes in the 2D scatter patterns are used to infer the location of the 90-deg scatter (polar angle). The 2D scatter patterns are shown to be symmetrical about the azimuth angle at 90 deg. Wide-angle comparisons between the experimental scatter patterns and Mie theory simulations are shown to be in good agreement. A method based on the Fourier transform analysis of the experimental and Mie simulation scatter patterns is developed for size differentiation.

Microscope-based label-free microfluidic cytometry.

A microscope-based label-free microfluidic cytometer capable of acquiring two dimensional light scatter patterns from single cells, pattern analysis of which determines cellular information such as cell size, orientation and inner nanostructure, was developed. Finite-difference time-domain numerical simulations compared favorably with experimental scatter patterns from micrometer-sized beads and cells. The device was capable of obtaining light scattering patterns from the smallest mature blood cells (platelets) and cord blood hematopoietic stem/progenitor cells 
(CD34 + cells) and myeloid precursor cells. The potential for evaluation of cells using this label-free microfluidic cytometric technique was discussed.

A miniaturized wide-angle 2D cytometer

We present an optical waveguide based cytometer that is capable of simultaneously collecting the light scattered by cells over a wide range of solid angles. Such comprehensive scattering data are a prerequisite for the microstructural characterization of cells.

Light scattering characterization of mitochondrial aggregation in single cells

Three dimensional finite-difference time-domain (FDTD) simulations are employed to show that light scattering techniques may be used to infer the mitochondrial distributions that exist within single biological cells. Two-parameter light scattering plots of the FDTD light scattering spectra show that the small angle forward scatter can be used to differentiate the case of a random distribution of mitochondria within a cell model from that in which the mitochondria are aggregated to the nuclear periphery. Fourier transforms of the wide angle side scatter spectra show a consistent highest dominant frequency, which may be used for size differentiation of biological cells with distributed mitochondria.

Label-free and noninvasive optical detection of the distribution of nanometer-size mitochondria in single cells

A microfluidic flow cytometric technique capable of obtaining information on nanometer-sized organelles in single cells in a label-free, noninvasive optical manner was developed. Experimental two-dimensional (2D) light scattering patterns from malignant lymphoid cells (Jurkat cell line) and normal hematopoietic stem cells (cord blood CD34+ cells) were compared with those obtained from finite-difference time-domain simulations. In the simulations, we assumed that the mitochondria were randomly distributed throughout a Jurkat cell, and aggregated in a CD34+ cell. Comparison of the experimental and simulated light scattering patterns led us to conclude that distinction from these two types of cells may be due to different mitochondrial distributions. This observation was confirmed by conventional confocal fluorescence microscopy. A method for potential cell discrimination was developed based on analysis of the 2D light scattering patterns. Potential clinical applications using mitochondria as intrinsic biological markers in single cells were discussed in terms of normal cells (CD34+ cell and lymphocytes) versus malignant cells (THP-1 and Jurkat cell lines).

Wide-angle light-scattering differentiation of organelle-size particle distributions in whole cells.

A finite‐difference time‐domain (FDTD) method is used to study the multiple scattering from many organelle‐size particles distributed in a biological cell. Conventional flow cytometry, where the small‐angle forward scatter (FSC) intensity and side scatter (SSC) intensity are used for cell characterizations, may have difficulties to differentiate the organelle distributions in biological cells. Based on the FDTD simulations, a light‐scattering methodology is proposed here to overcome such a problem. This method differentiates the dense and sparse distributions of organelle‐size particles in a cell, by counting the peak numbers in both large‐angle FSC and wide‐angle SSC, with the multiple scattering effects being considered. Implemented with a wide‐angle microfluidic cytometer, the approach demonstrated in this theoretical study may find potential applications in clinics for label‐free cell physiological study.

2D light scattering static cytometry for label-free single cell analysis with submicron resolution.

Conventional optical cytometric techniques usually measure fluorescence or scattering signals at fixed angles from flowing cells in a liquid stream. Here we develop a novel cytometer that employs a scanning optical fiber to illuminate single static cells on a glass slide, which requires neither microfluidic fabrication nor flow control. This static cytometric technique measures two dimensional (2D) light scattering patterns via a small numerical aperture (0.25) microscope objective for label‐free single cell analysis. Good agreement is obtained between the yeast cell experimental and Mie theory simulated patterns. It is demonstrated that the static cytometer with a microscope objective of a low resolution around 1.30 μm has the potential to perform high resolution analysis on yeast cells with distributed sizes. The capability of the static cytometer for size determination with submicron resolution is validated via measurements on standard microspheres with mean diameters of 3.87 and 4.19 μm. Our 2D light scattering static cytometric technique may provide an easy‐to‐use, label‐free, and flow‐free method for single cell diagnostics.

Rapid simulation of wide-angle scattering from mitochondria in single cells

It has been shown that the mitochondria are the dominant source of large-angle light scattering from human cells. In the limit of small mitochondria, we show that the large-angle (isotropic) light scattering of mitochondria may be analyzed and simulated with an adaptation of classical X-ray diffraction theory. In addition, we show that this approach may be extended to the case of anisotropic scatter. These results enable the rapid simulation and analysis of mitochondrial scattering patterns and allow the determination of some aspects of cell structure directly from experimental scattering patterns.

Exosomes derived from human umbilical cord mesenchymal stem cells protect against cisplatin-induced ovarian granulosa cell stress and apoptosis in vitro

Human umbilical cord mesenchymal stem cells (huMSCs) can treat primary ovarian insufficiency (POI) related to ovarian granulosa cell (OGC) apoptosis caused by cisplatin chemotherapy. Exosomes are a class of membranous vesicles with diameters of 30–200 nm that are constitutively released by eukaryotic cells. Exosomes mediate local cell-to-cell communication by transferring microRNAs and proteins. In the present study, we demonstrated the effects of exosomes derived from huMSCs (huMSC-EXOs) on a cisplatin-induced OGC model in vitro and discussed the preliminary mechanisms involved in these effects. We successfully extracted huMSC-EXOs from huMSC culture supernatant and observed the effective uptake of exosomes by cells with fluorescent staining. Using flow cytometry (with annexin-V/PI labelling), we found that huMSC-EXOs increased the number of living cells. Western blotting showed that the expression of Bcl-2 and caspase-3 were upregulated, whilst the expression of Bax, cleaved caspase-3 and cleaved PARP were downregulated to protect OGCs. These results suggest that huMSC-EXOs can be used to prevent and treat chemotherapy-induced OGC apoptosis in vitro. Therefore, this work provides insight and further evidence of stem cell function and indicates that huMSC-EXOs protect OGCs from cisplatin-induced injury in vitro.

Pattern recognition cytometry for label-free cell classification by 2D light scattering measurements

We develop a pattern recognition cytometric technique for label-free cell classification. Two dimensional (2D) light scattering patterns from single cells and cell aggregates are obtained with a static cytometer. Good performance of the cytometric setup is verified by comparing yeast cell experimental results with theoretical simulations. Adaptive boosting (AdaBoost) method (a machine learning algorithm) is adopted for the analysis of the 2D light scattering patterns. It is shown that aggregates of three yeast cells can be well differentiated from aggregates of four yeast cells by this pattern recognition cytometric technique. We demonstrate that the pattern recognition cytometry can perform label-free classification of normal cervical cells and HeLa cells with a high accuracy rate.