As a biomarker for the diagnosis and treatment monitoring of various diseases, exosomes widely exist in body fluids such as blood, urine and saliva. However, its small particle size and low content are difficult to enrich. Therefore, a fast, high-purity enrichment method, and a fast, high-sensitivity, high-resolution, and low detection limit detection method are particularly important. We developed an automated fully integrated system for the enrichment and detection of exosomes. Using magnetic beads modified with anionic polymers to capture exosomes, adjust the pH of the exosome solution to acidic, and use the electrostatic adsorption between the positive charge on the surface of exosomes and the negative charge on the surface of the anionic polymer-modified substrate to achieve exosome capture. The captured extracellular vesicles are eluted from the surface of the magnetic beads by using a neutral or slightly alkaline eluent and using electrostatic repulsion to achieve the purpose of separation and enrichment of extracellular vesicles. The method is fast and efficient, can be automated with a small instrument, and can exclude favorable nucleic acid interferences. The eluted exosome protein is fixed on the substrate by chemical modification using quantitative interference exosome surface protein detection technology, and the connection between the exosome surface protein and the antibody is realized by immunoadsorption. Hyperspectral interferometry was used to quantitatively analyze the optical path increment on the substrate surface, to determine whether the exosome sample was bound to the antibody, and to detect the protein content of the exosome surface in parallel. This method can achieve sub-nanometer detection accuracy, and can detect exosomes whose size is smaller than the diffraction limit. Finally, the enrichment and detection of exosomes were automated.
Quantitative phase imaging (QPI) has quickly emerged as a powerful tool for label-free living cell morphology and metabolism monitoring. However, for current QPI techniques, interference signals from different layers overlay with each other and impede nanoscale optical sectioning. This phenomenon leads to unsatisfactory performances for optically thick or complex scattering biological samples. To address this challenge, we have developed an alternative quantitative phase microscopy with computational hyperspectral interferometry. Nanoscale optical sectioning could be achieved with Fourier domain spectral decomposition. Morphological fluctuations and refractive index distribution could be reconstructed simultaneously with 89.2 nm axial resolution and 1.91 nm optical path difference sensitivity. With this method, we established a label-free cell imaging system for long-term cellular dry mass measurement and in-situ dynamic single cell monitoring. Different intrinsic cell growth characteristics of dry mass between HeLa cells and Human Cervical Epithelial Cells (HCerEpiC) were studied. The dry mass of HeLa cells consistently increased before M phase, whereas that of HCerEpiC increased and then decreased. The maximum growth rate of HeLa cells was 11.7% higher than that of HCerEpiC. We also use the proposed method and system to explore the relationship between cellular dry mass distributions and drug effects for cancer cells. The results show that cells with higher nuclear dry mass and nuclear density standard deviations were more likely to survive the chemotherapy. The presented work shows potential values for cell growth dynamics research, cell health characterization, medication guidance and adjuvant drug development.
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