Nanodiamonds with color centers are to an increasing degree investigated as intracellular biosensors for magnetic fields, electrical fields, or temperature changes, as well as for abundance of free radicals or pH inside live cells. A common color-center is the nitrogen-vacancy (NV) center, where a substitutional nitrogen atom is positioned next to a vacancy within the diamond host crystal. The nanodiamond with negatively charged NV− center is particularly versatile due to its biocompatibility and its purely optical addressability. Our work aims to use NV-center nanodiamonds both as intracellular biosensors and as probe particles within an optical trap to determine the viscoelastic properties of the intracellular environment in single-cell studies. For this to be successful, several prior steps are needed: 1) The uptake of nanodiamonds within the cells should be characterized, including studies of subcellular localization, and a controllable protocol developed; 2) Any effect of the trapping laser on the NV-center sensing should be characterized and understood, and a protocol for stable trapping along with accurate biosensing should be developed. In this work we summarize the preliminary findings of our ongoing investigations to address these points. We show results of T1-relaxometry with and without CW NIR laser irradiation in a suspension cell model, analyze optical trapping of nanodiamonds with CW light in an adhesion cell model, and investigate implications of the presence of the optical trapping laser on T1-relaxometry measurements.
Flow cytometry (FC) is a pivotal tool for studying the physical and chemical properties of particles. State-of-the-art FC systems are highly advanced, yet they are expensive, bulky, and require high sample volume, qualified operators, and periodic maintenance. The manipulation of particles suspended in viscoelastic fluids has received increasing attention, especially for miniaturized flow cytometry technologies. This study presents a miniaturized optical capillary FC device using the viscoelastic focusing technique. A straight, one inlet/outlet microcapillary device is precisely aligned to a fiber-coupled laser source and detectors. Forward scattered, side scattered, and fluorescently emitted light signals are collected and analyzed in a real-time environment. The developed platform fits onto an inverted microscope stage enabling real-time microscopy imaging of the particles of interest together with the flow cytometry analysis. We achieved stable viscoelastic focusing and performed FC measurements for rigid polystyrene beads (diameters: 2 – 15 μm), non-spherical human erythrocytes, and canonical shape metaphase human chromosomes. We performed cytometry measurements with a throughput of 100 events/s yielding a coefficient of variation of 2%. This newly developed FC device is a versatile tool and can be operated with any inverted microscope to get the mutual benefits of optical and imaging FC measurements. Furthermore, it is possible to extend these benefits by adding more back-end tools, such as optical trapping and Raman spectroscopy.
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