研究目的
To develop and validate Scatter Enhanced Phase Contrast (SEPC) microscopy as a label-free method for monitoring the uptake and transport dynamics of engineered nanomaterials (ENMs) in living cells, addressing the limitations of fluorescent labels and enabling the study of various metal and metal oxide nanoparticles.
研究成果
SEPC microscopy is a powerful, label-free tool for studying nanoparticle uptake and transport in living cells, revealing spatial heterogeneities and distinct transport modes. It enables the rational exploration of nanomaterial surface properties and their roles in endocytosis, with implications for drug delivery and environmental risk assessment. Future work could extend to polymeric materials and long-term fate studies of intracellular particles.
研究不足
The technique is limited to materials with sufficient optical scattering properties, as predicted by Lorenz-Mie theory (e.g., SiO2 nanoparticles show poor scattering and are difficult to detect below ~80 nm). It may not be effective for non-adherent spherical cells due to membrane curvature issues. The method relies on aggregation for smaller nanoparticles, and real-time tracking is constrained by equipment capabilities such as exposure times and condenser working distances.
1：Experimental Design and Method Selection:
The study uses SEPC microscopy, combining darkfield and phase contrast techniques with Lorenz-Mie theory for predicting nanoparticle scattering. It involves real-time imaging of nanoparticle-cell interactions.
2：Sample Selection and Data Sources:
Human umbilical vascular endothelial cells (HUVECs) and human aortic smooth muscle cells (HASMCs) are used, exposed to nanoparticles such as Au, Ag, TiO2, CeO2, Al2O3, Fe2O3, and SiO2. Nanoparticles are synthesized and characterized as per previous reports.
3：Nanoparticles are synthesized and characterized as per previous reports.
List of Experimental Equipment and Materials:
3. List of Experimental Equipment and Materials: Includes an Olympus ix83 inverted microscope with phase contrast plate, oblique angle LED ring illuminator (Amscope LED-144-YK), LUCPlanFLNPh objectives (20x and 40x), Hamamatsu Orca Flash 4.0 C11440 camera, thermal heater (Air-Therm ATX-H), and stage top incubator. Nanoparticles are prepared in suspensions (e.g., 10 μg/mL).
4：0 C11440 camera, thermal heater (Air-Therm ATX-H), and stage top incubator. Nanoparticles are prepared in suspensions (e.g., 10 μg/mL).
Experimental Procedures and Operational Workflow:
4. Experimental Procedures and Operational Workflow: Cells are cultured and exposed to nanoparticles. SEPC imaging is performed at intervals (e.g., 2-4 seconds for dynamics, 20 minutes for uptake studies). Hyperspectral imaging and Raman spectroscopy are used for nanoparticle identity confirmation. Microcontact printing with PDMS stamps creates reproducible cell morphologies for normalized studies.
5：Data Analysis Methods:
Particle tracking involves measuring velocity, mean squared displacement (MSD), and diffusivity exponent (α). Data is analyzed using linear least squares fitting for MSD plots, and composite images are created for ensemble dynamics. Statistical analysis includes signal-to-noise ratio calculations and intensity measurements.
独家科研数据包，助您复现前沿成果，加速创新突破
获取完整内容
