Three-dimensional super-resolution correlation-differential confocal microscopy with nanometer axial focusing accuracy

We present a correlation-differential confocal microscopy (CDCM), a novel method that can simultaneously improve the three-dimensional spatial resolution and axial focusing accuracy of confocal microscopy (CM). CDCM divides the CM imaging light path into two paths, where the detectors are before and after the focus with an equal axial offset in opposite directions. Then, the light intensity signals received from the two paths are processed by the correlation product and differential subtraction to improve the CM spatial resolution and axial focusing accuracy, respectively. Theoretical analyses and preliminary experiments indicate that, for the excitation wavelength of λ = 405 nm, numerical aperture of NA = 0.95, and the normalized axial offset of uM = 5.21, the CDCM resolution is improved by more than 20% and more than 30% in the lateral and axial directions, respectively, compared with that of the CM. Also, the axial focusing resolution important for the imaging of sample surface profiles is improved to 1 nm

Improving spatial resolution of confocal Raman microscopy by super-resolution image restoration

A new super-resolution image restoration confocal Raman microscopy method (SRIR-RAMAN) is proposed for improving the spatial resolution of confocal Raman microscopy. This method can recover the lost high spatial frequency of the confocal Raman microscopy by using Poisson-MAP super-resolution imaging restoration, thereby improving the spatial resolution of confocal Raman microscopy and realizing its super-resolution imaging. Simulation analyses and experimental results indicate that the spatial resolution of SRIR-RAMAN can be improved by 65% to achieve 200 nm with the same confocal Raman microscopy system. This method can provide a new tool for high spatial resolution micro-probe structure detection in physical chemistry, materials science, biomedical science and other areas

Confocal Raman image method with maximum likelihood method

With the increasing interest in nano microscopic area, such as DNA sequencing, micro structure detection of molecular nano devices, a higher requirement for the spatial resolution of Raman spectroscopy is demanded. However, because of the weak Raman signal, the pinhole size of confocal Raman microscopy is usually a few hundreds microns to ensure a relatively higher spectrum throughput, but the large pinhole size limits the improvements of spatial resolution of confoal Raman spectroscopy. As a result, the convential confocal Raman spectroscopy has been unable to meet the needs of science development. Therefore, a confocal Raman image method with Maximum Likelihood image restoration algorithm based on the convential confocal Raman microscope is propose. This method combines super-resolution image restoration technology and confocal Raman microscopy to realize super-resolution imaging, by using Maximum Likelihood image restoration algorithm based on Poisson-Markov model to conduct image restoration processing on the Raman image, and the high frequency information of the image is recovered, and then the spatial resolution of Raman image is improved and the super-resolution image is realized. Simulation analyses and experimental results indicate that the proposed confocal Raman image method with Maximum Likelihood image restoration algorithm can improve the spatial resolution to 200 nm without losing any Raman spectral signal under the same condition with convential confocal Raman microscopy, moreover it has strong noise suppression capability. In conclusion, the method can provide a new approach for material science, life sciences, biomedicine and other frontiers areas. This method is an effective confocal Raman image method with high spatial resolution

Synchronous nanoscale topographic and chemical mapping by differential-confocal controlled Raman microscopy

Confocal Raman microscopy is currently used for label-free optical sensing and imaging within the biological, engineering, and physical sciences as well as in industry. However, currently these methods have limitations, including their low spatial resolution and poor focus stability, that restrict the breadth of new applications. This paper now introduces differential-confocal controlled Raman microscopy as a technique that fuses differential confocal microscopy and Raman spectroscopy, enabling the point-to-point collection of three-dimensional nanoscale topographic information with the simultaneous reconstruction of corresponding chemical information. The microscope collects the scattered Raman light together with the Rayleigh light, both as Rayleigh scattered and reflected light (these are normally filtered out in conventional confocal Raman systems). Inherent in the design of the instrument is a significant improvement in the axial focusing resolution of topographical features in the image (to ∼1 nm ), which, when coupled with super-resolution image restoration, gives a lateral resolution of 220 nm. By using differential confocal imaging for controlling the Raman imaging, the system presents a significant enhancement of the focusing and measurement accuracy, precision, and stability (with an antidrift capability), mitigating against both thermal and vibrational artefacts. We also demonstrate an improved scan speed, arising as a consequence of the nonaxial scanning mode

Three-dimensional resolution-enhancement divided aperture correlation-differential confocal microscopy with nanometer axial focusing capability

Divided aperture confocal microscopy (DACM) provides an improved imaging depth, imaging contrast, and working distance at the expense of spatial resolution. Here, we present a new method-divided aperture correlation-differential confocal microscopy (DACDCM) to improve the DACM resolution and the focusing capability, without changing the DACM configuration. DACDCM divides the DACM image spot into two round regions symmetrical about the optical axis. Then the light intensity signals received simultaneously from two round regions by a charge-coupled device (CCD) are processed by correlation manipulation and differential subtraction to improve the DACM spatial resolution and axial focusing capability, respectively. Theoretical analysis and preliminary experiments indicate that, for the excitation wavelength of λ = 632.8 nm, numerical aperture NA = 0.8, and normalized offset vM = 3.2 of the two regions, the DACDCM resolution is improved by 32.5% and 43.1% in the x and z directions, simultaneously, compared with that of the DACM. The axial focusing resolution used for the sample surface profile imaging was also significantly improved to 2 nm

Confocal Raman spectroscopy method based on quadratic curve fitting

Raman spectroscopy plays an improtant role in analytical science because of its unique characteristics, such as non-contact and non-destructive detecting, fewer sample consumption, high sensitivity and other characteristics, and it provides a powerful analytical tool for the modern basic research fields. Because of the combination of confocal microscopy technology and Raman spectroscopy technology, confocal Raman microscopy has the advantage of both high resolution spectroscopy and chromatography detection, which inherits from confocal microscopy and raman spectroscopy. As a result, it is widely used in many fields, such as physical chemistry, materials science, biomedical, archaeological, cultural identification, and forensic science. But with the environmental changing, the system drifting or other issues, during the long detection process, the system turns to defocusing. As a result, during the hole scanning process, the system can not be focused on every detection point, and then it would lead to a mistake. Eventurally, conventional confocal Raman system could obtain the presence of measurement error even erroneous results in the long process. In this paper, on the basis of conventional confocal Raman system, a confocal Raman spectroscopy method based on quadratic curve fitting is proposed to solve this problem. Based on the principle that the maxium of the concal curve corresponding the system foucs, the steps to find system foucus as follows: fist, usesing quadratic curve to fit confocal curve; second, finding the maxium of the confocal curve; and last obtaining the system foucs. With this method, during the scanning process, every point should be focused, therefore, the effect of defocusing is eliminated efficiently, and accurate measurements of confocal Raman spectroscopy system is achieved.Through simulation and experimental results show that: the proposed method that confocal Raman spectroscopy method based on quadratic curve fitting can effectively eliminate the influence of system defocus on experimental result, and effectively improve the axial system of fixed focus accuracy, which could provide a guarantee for further application of confocal Raman spectroscopy. This anti-drift method is effective and accurate in focusing with great potential to be applied in broader areas

Moringa oleifera leaf alleviates functional constipation via regulating the gut microbiota and the enteric nervous system in mice

Moringa oleifera Lam. leaf is not only a new food resource in China, but also a traditional medicinal plant. It is commonly used in the folk to alleviate constipation, but its laxative mechanism is not fully understood. Hence we investigated it in loperamide-induced functional constipation (FC) mice. The results showed that MOAE significantly regulated not only gastrointestinal hormones and neurotransmitters in serum but also important gastrointestinal motility factors in the enteric nervous system (ENS)-interstitial cells of Cajal (ICCs)-smooth muscle cell (SMC) network. Meanwhile, MOAE attenuated intestinal inflammation, increased cecal short-chain fatty acid levels and colonic antimicrobial peptide expression, and improved the impaired intestinal barrier function in loperamide-induced FC mice. In addition, MOAE also increased fecal water content by inhibiting the mRNA expression of colonic aquaporins (Aqp3 and Aqp4) in FC mice. Interestingly and importantly, MOAE affected the intestinal microbiota by inhibiting some key “constipation-causing” microbiota, such as Bacteroidaceae, Clostridiaceae, Bacteroides, and Ruminococcus, and promoting the growth of other important “constipation-curing” microbiota, such as Butyricoccus, Tyzzerella, and Desulfovibrio. These important taxa are significantly associated with a variety of indicators of constipation. These findings suggest that MOAE can promote defecation through its rich chemical composition to modulate the ENS-ICCs-SMCs network and the gut microecosystem

The Lid Domain in Lipases: Structural and Functional Determinant of Enzymatic Properties

Lipases are important industrial enzymes. Most of the lipases operate at lipid–water interfaces enabled by a mobile lid domain located over the active site. Lid protects the active site and hence responsible for catalytic activity. In pure aqueous media, the lid is predominantly closed, whereas in the presence of a hydrophobic layer, it is partially opened. Hence, the lid controls the enzyme activity. In the present review, we have classified lipases into different groups based on the structure of lid domains. It has been observed that thermostable lipases contain larger lid domains with two or more helices, whereas mesophilic lipases tend to have smaller lids in the form of a loop or a helix. Recent developments in lipase engineering addressing the lid regions are critically reviewed here. After on, the dramatic changes in substrate selectivity, activity, and thermostability have been reported. Furthermore, improved computational models can now rationalize these observations by relating it to the mobility of the lid domain. In this contribution, we summarized and critically evaluated the most recent developments in experimental and computational research on lipase lids

High spatial resolution biaxial differential confocalspectrum microscopic imaging method and apparatus

The invention belongs to the technical field of spectrum measurement, and relates to a high spatial resolution biaxial differential confocal spectrum imaging method and an apparatus. According to the present invention, the biaxial differential confocal microscopy technology and the spectrum detection technology are fused, focal spot cutting differential detection is adopted so as to achieve precise imaging of the geometry position, simplify the optical path structure of the traditional differential confocal microscopy system, inherit the advantages of large visual field and large work distance of the biaxial microscopy technology, and achieve high spatial resolution spectrum integrated detection of the system; and high spatial resolution is provided, three modes such as three-dimensional tomography geometry imaging, spectrum detection and micro-region spectrum tomography imaging are provided, a new solving approach is provided for micro-region spectrum detection, and broad application prospects are provided in the fields of biomedicine, physical material science and the like

High-space-resolution double-shaft confocal atlas micro-imaging method and device

The invention belongs to the technical field of spectral measurement and relates to a high-space-resolution double-shaft confocal atlas micro-imaging method and device. According to a core concept of the method, a double-shaft confocal micro-technique and a spectral detection technique are organically merged, and a normally-forgotten Reyleigh light is utilized to carry out auxiliary detection, so that the space resolution of a system is improved, and three detection modes including three-dimensional tomographic imaging, spectral detection and microcell atlas tomographic imaging are realized. The method and the device have the advantages of high space resolution, accuracy in positioning, high spectral detection sensitivity and the like, have wide application prospects in the fields of biomedicines, physical material science, petrochemical engineering, environmental science and the like and provide new ways for the high-space-resolution detection of microcell three-dimensional geometric positions and spectra