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.
Label free point mutation detection is particularly momentous in the area of biomedical research and clinical diagnosis since gene mutations naturally occur and bring about highly fatal diseases. In this paper, a label free and high sensitive approach is proposed for point mutation detection based on hyperspectral interferometry. A hybridization strategy is designed to discriminate a single-base substitution with sequence-specific DNA ligase. Double-strand structures will take place only if added oligonucleotides are perfectly paired to the probe sequence. The proposed approach takes full use of the inherent conformation of double-strand DNA molecules on the substrate and a spectrum analysis method is established to point out the sub-nanoscale thickness variation, which benefits to high sensitive mutation detection. The limit of detection reach 4pg/mm2 according to the experimental result. A lung cancer gene point mutation was demonstrated, proving the high selectivity and multiplex analysis capability of the proposed biosensor.
Optical microscopy technology has achieved great improvements in the 20th century. The detection limit has reached about twenty nanometers (with near-field optics, STED, PALM and STORM). But in the application areas such as life science, medical science, clinical treatment and especially in vivo dynamic measurement, mutual restrictions still exist between numeric aperture/magnification and working distance, fluorescent dependent, and between resolution and frame rate/field size, etc. This paper explores a hyperspectral scanning super-resolution label free molecules imaging method based on the white light interferometry. The vertical detection resolution was approximate to 1 nm which is the thickness of a single molecular layer and dynamic measuring range of thickness reaches to 10 μm. The spectrum-shifting algorithm is developed for robust restructure of images when the pixels are overlapped. Micro-biochip with protein binding and DNA amplification could be detected by using this spectral scanning super-resolution molecules imaging in label free. This method has several advantages as following: Firstly, the decoding and detecting steps are combined into one step. It makes tests faster and easier. Secondly, we used thickness-coded, minimized chips instead of a large microarray chip to carry the probes. This accelerates the interaction of the biomolecules. Thirdly, since only one kind of probes are attached to our thickness-coded, minimized chip, users can only pick out the probes they are interested in for a test without wasting unnecessary probes and chips.
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