Super-resolution fluorescence polarization microscopy

Fluorescence polarization is related to the dipole orientation of chromophores,making fuores-cence polarization microscopy possible to_reveal structures and functions of tagged cellularorganelles and biological macromolecules.Several recent super resolution techniques have beenapplied to fluorescence polarization microscopy,achieving dipole measurement at nanoscale.In this review,we summarize both difraction limited and super resolution fluorescence polari-zation microscopy techniques,as well as their applications in biological imaging.

Advancing respirable coal mine dust source apportionment:a preliminary laboratory exploration of optical microscopy as a novel monitoring tool

Exposure to respirable coal mine dust(RCMD)can cause chronic and debilitating lung diseases.Real-time monitoring capabilities are sought which can enable a better understanding of dust components and sources.In many underground mines,RCMD includes three primary components which can be loosely associated with three major dust sources:coal dust from the coal seam itself,silicates from the surrounding rock strata,and carbonates from the inert‘rock dust’products that are applied to mitigate explosion hazards.A monitor which can reliably partition RCMD between these three components could thus allow source apportionment.And tracking silicates,specifically,could be valuable since the most serious health risks are typically associated with this component-particularly if abundant in crystalline silica.Envisioning a monitoring concept based on field microscopy,and following up on prior research using polarized light,the aim of the current study was to build and test a model to classify respirable-sized particles as either coal,silicates,or carbonates.For model development,composite dust samples were generated in the laboratory by successively depositing dust from high-purity materials onto a sticky transparent substrate,and imaging after each deposition event such that the identity of each particle was known a priori.Model testing followed a similar approach,except that real geologic materials were used as the source for each dust component.Results showed that the model had an overall accuracy of 86.5%,indicating that a field-microscopy based moni-tor could support RCMD source apportionment and silicates tracking in some coal mines.

Erythropoietic Protoporphyria (A Light, Polarization and Electron Microscopical Study of the Liver in One Patient)

Objective\ To explore the hepatic pathology of a patient with erythropoietic protoporphyria (EPP). Methods\ Percutaneous liver biopsy was performed with a Chiba needle in a 31 year old man suffering from EPP. The sample was fixed in 10% formalin solution, and the paraffin embedded section was stained with H E, PAS, etc. Unstained paraffin embedded and H E stained paraffin embedded sections were examined under polarization microscope. Ultrathin sections were examined in a transmission electron microscope. Results\ In H E stained sections, deposits of dark reddish brown pigment were seen in the hepatocytes, Kupffer cells, portal macrophages and plugs in the lumen of bile canaliculi and ducts. Under light microscope, such deposits, with rare exception, exhibited striking birefringence with the unique shape of 'Maltese cross'. Non membrane limited compact masses of crystals were straight or slightly curved and their dimensions (40 640 nm in length and 6 22 nm in width) were different under the transmission electron microscope. Conclusion\ Microscopy, especially polarization microscopy, provides a highly sensitive and specific technique for the diagnosis of EPP.

New method for fast morphological characterization of organic polycrystalline films by polarized optical microscopy

A new method to visualize the large-scale crystal grain morphology of organic polycrystalline films is proposed. First,optical anisotropic transmittance images of polycrystalline zinc phthalocyanine（Zn Pc） films vacuum deposited by weak epitaxial growth（WEG） method were acquired with polarized optical microscopy（POM）. Then morphology properties including crystal grain size, distribution, relative orientation, and crystallinity were derived from these images by fitting with a transition dipole model. At last, atomic force microscopy（AFM） imaging was carried out to confirm the fitting and serve as absolute references. This method can be readily generalized to other organic polycrystalline films, thus providing an efficient way to access the large-scale morphologic properties of organic polycrystalline films, which may prove to be useful in industry as a film quality monitoring method.

Quantification of polarization distribution from polarizing light microscopy images of ferroelectric single crystals

Qualification of polarization can be realized either on a macroscopic scale as an average property by P-E hysteresis measurements or on a nano/micro scale by piezoelectric force microscopy,transmission electron microscopy,scanning electron microscopy,and so on.However,visualization and qualification of polarization distribution in the micron to millimeter scale is still a challenge.Polarizing light microscopy(PLM)is often used in the study of ferroelectric domain structures mainly for domain patterns.A phe-nomenon called“chromatic polarization”has been observed in transparent ferroelectric crystals by using a crossed-PLM system viewed with white light,which contains rich information about local polariza-tion distribution.In this study,an automatic full-angle light intensity detection(AFALID)algorithm com-bined with colorimetry is developed to analyze the distribution of nonuniform local spontaneous polar-ization distribution in transparent ferroelectric single crystals.Temperature-dependent spontaneous po-larizations from the color analysis for PMN-0.36PT single crystals with single tetragonal domain state are in good coincidence with those extracted from temperature-dependent hysteresis loops and pyroelectric current measurements.We further apply this method to quantify the nonuniform domain distributions with nano-indentations.This non-contact and non-destructive characterization can provide fast and au-tomatic detection of polarization distributions in ferroelectric materials.

Investigation of Gd_(3)Ga_(5)O_(12) by Micropolarimetry

This paper summarizes the results of investigation of garnets Gd3Ga5O12 (GGG) on the basis of their optical activity. Dispersion of Verdet constant was measured in the visible light range and in a strong magnetic field. The applied technique, namely micropolarimetry, exhibits high resolution which corresponds to the type of polarization microscope used in the investigation.

MORPHOLOGICAL AND KINETIC STUDIES OF PHASE TRANSITIONS OF A SIDE-CHAIN LIQUID CRYSTALLINE POLYMER

The morphological changes of a side-chain liquid-crystalline polymethacrylate during isotropization and liquid-crystallization transitions were studied by means of polarizing microscopy. These transitions were found to be composed of the initiation of a new phase at local places of the old phase matrix and the growth of the new phase: domains. The kinetics of the liquid-crystallization of the polymer from an isotropic melt to a smectic mesophase was also investigated. The isothermal process of the transition can be described by the Avrami equation. The values of the Avrami exponent were found to be around 2.6. which is lower than the value usually obtained for crystallization transition of polymers, but larger than that reported for liquid-crystallization transition of main-chain polymers. These results may indicate the difference in growth geometry of new phase during transition between crystallization and liquid-crystallization in general and between liquid-crystallization of main-chain and side-chain polymers. It was found that the liquid-crystallization of the used side-chain polymethacrylate may occur at small undercoolings with high transformation rate similar to that of main-chain polymers and small-molecule liquid crystals, while the crystallization of polymers can only proceed at large undercoolings. These phenomena can be explained by the idea that the surface free energy of nucleus during liquid-crystallization transition is less than that for crystallization, and evidence was obtained from analysis of the temperature dependence of the transformation rate.

STUDIES ON THE MORPHOLOGY OF THERMOTROPIC POLYESTER DURING TRANSITIONS BETWEEN ISOTROPIC AND ANISOTROPIC MELTS

The morphology of a thermotropic aromatic polyester during the transitions between isotropic and anisotropic melts was studied by means of polarizing microscopy and small angle light scattering techniques. Both transition processes were found to be composed of the initiation of a new phase at local places of the old phase matrix and the growth of the new phase domains. A change of Hv scattering patterns was observed in the transition temperature ranges and was explained as the result of heterogeneity of the mesophase system, but not the change in ordering of a homogeneous mesophase.

MORPHOLOGICAL STUDIES OF A THERMOTROPIC SIDE-CHAIN LIQUID CRYSTALLINE POLYMER DURING MESOPHASE TRANSITIONS

The morphological features of a side-chain liquid crystalline polymer during the mesophase transitions were investigated by using the DSC technique. The polymer used was polyacrylate with mesogens of three benzene rings attached to the main chain through a flexible spacer. A special two-phase texture was observed in the transition temperature range. Similar to main-chain liquid crystalline polymers the transition process of the side-chain liquid crystalline polymer was composed of an initiation of the new phase at local places of the old phase matrix and a growth process of the new phase domains.

Mesomorphic Studies on a Series of Nickel (II) Centered Aromatic Complexes Displaying Liquid Crystalline Phases

Two new series (A & B) of three materials each based on the aroyl hydrazinato-nickel (II) complex were synthesized and characterized. The core molecule in these series consists of two 4-benzoyloxy benzylidene moieties and two benzene rings attached at the azomethine moiety. These latter benzene rings have one or two alkoxy chains comprised of either 10, 12, 16 or 18 carbon atoms. The characterization of these two series by polarized optical microscopy and differential scanning calorimetry is described herein. Upon cooling from the Isotropic phase, three of the six materials display a monophasic columnar phase and the other three possess a biphasic nematic and columnar phases. Upon heating, all six materials have a clearance point at high temperatures without displaying mesomorphic behavior. In series B mesogens, it was observed that the longer the hydrocarbon tail length, the lower the clearance point.

Phase Transition Investigations of a Series of Aromatic Naphthalene-2-yl-4-(Alkoxy) Benzoate and Naphthalene-1-yl-4-(Alkoxy) Benzoate Materials

The mesogenic properties of a homologous series of aromatic ester materials are investigated. Single tail and double tail materials were synthesized to compare with other series we prepared and published earlier. The phase sequences and transition temperatures were obtained by polarized light microscopy and differential scanning calorimetry. Only the single tail materials with the ester attached to naphthalene at position 2 display mesogenicbehavior. Two materials (ZH 29 & ZH 32) possess a monophasic nematic phase and another two materials (ZH 14 & ZH 35) display biphasic nematic and smectic (A) phases.

Application of polarized light microscopy on microstructural characterization of semisolid slurry

In this paper,two ways of micro structural characterization,optical microscopy(OM) and polarized light microscopy(PLM),were both employed to describe the micro structure of semisolid slurry prepared by swirling enthalpy equilibration device(SEED).The results show that PLM is more reliable and accurate than OM to describe the special morphology feature of semisolid slurry made by SEED process.Meanwhile,the effects of pouring temperature and mass of molten liquid on the primary α-Al particle size and morphology were also investigated using PLM.The quantitative metallographic results measured from PLM demonstrate that the grain size and morphology and their distribution are significantly affected by both pouring temperature and the mass of molten liquid.The grain size poured with 2.7 kg liquid decreases from 659 to186 μm,and grain morphology transforms from dendrite to globular structure with pouring temperature reducing from690 to 630℃.The decreasing pouring temperature also promotes the distribution of spherical structure on the cross section.Meanwhile,the mass of molten liquid decreasing from 2.7 to 2.3 kg can decrease the grain size by maximum of 44% at high pouring temperature.

Super-resolution dipole orientation mapping via polarization demodulation

Fluorescence polarization microscopy(FPM)aims to detect the dipole orientation of fluorophores and to resolve structural information for labeled organelles via wide-field or confocal microscopy.Conventional FPM often suffers from the presence of a large number of molecules within the diffraction-limited volume,with averaged fluorescence polarization collected from a group of dipoles with different orientations.Here,we apply sparse deconvolution and least-squares estimation to fluorescence polarization modulation data and demonstrate a super-resolution dipole orientation mapping(SDOM)method that resolves the effective dipole orientation from a much smaller number of fluorescent molecules within a sub-diffraction focal area.We further apply this method to resolve structural details in both fixed and live cells.For the first time,we show that different borders of a dendritic spine neck exhibit a heterogeneous distribution of dipole orientation.Furthermore,we illustrate that the dipole is always perpendicular to the direction of actin filaments in mammalian kidney cells and radially distributed in the hourglass structure of the septin protein under specific labelling.The accuracy of the dipole orientation can be further mapped using the orientation uniform factor,which shows the superiority of SDOM compared with its wide-field counterpart as the number of molecules is decreased within the smaller focal area.Using the inherent feature of the orientation dipole,the SDOM technique,with its fast imaging speed(at sub-second scale),can be applied to a broad range of fluorescently labeled biological systems to simultaneously resolve the valuable dipole orientation information with super-resolution imaging.

Background-free three-dimensional selective imaging of anisotropic plasmonic nanoparticles

There is an increasing demand for advanced optical imaging techniques that can detect and resolve nanosize objects at a spatial resolution below the optical diffraction limit, especially in three-dimensional （3D） cellular environments. In this study, using a polarization-activated localization scheme based on the orientation-dependent properties of anisotropic plasmonic metal nanoparticles （MNPs）, ＂photoswitchable＂ imaging of single gold nanorods （AuNRs） was accomplished not only in two dimensions but also in three dimensions. Moreover, the Rayleigh scattering background arising from the congested subcellular structures was efficiently suppressed. Thus, we obtained the 3D distributions of both the position and the orientation of the AuNRs inside the cells and investigated their intemalization kinetics. To our knowledge, this is the first demonstration of the confocal-like 3D imaging of non-fluorescence nanoparticles with a high resolution and almost zero background. This technique is easy to implement and should greatly facilitate MNP studies and applications in biomedicine and biology.

Preparation and Characterization of Mesophase Pitch via Co-Carbonization of Waste Polyethylene/Petroleum Pitch

The low-cost petroleum pitch and waste polyethylene （WPE） were used as raw materials to prepare the mesophase pitch by co-carbonization method and the forming mechanization of mesophase pitch was also investigated. Polarized microscopy, softening point, Fourier transform infrared spectroscopy （FTIR） and thermogravimetric analysis （TGA） were used to characterize and analyze the properties and struc- ture of the mesophase pitch. The results showed that the carbonization yield of the modified pitch was high when 1-2 wt% WPE was added and the property of mesophase pitch （MP1-450-4 and MP2-450-4） prepared by thermal polymerization was excellent. Moreover, when the treatment temperature was above 420 ~C, the mesophase development of the modified pitch may be entire and 100% streamline texture mesophase can form. During the co-carbonization of WPEJpetroleum pitch, a large number of naphthenic structures and methylene bridges may be generated, which can improve the properties of the obtained mesophase pitch.

Morphological and microscopic characterization of five commonly-used testacean traditional Chinese medicines

Testacean traditional Chinese medicine(TTCM), derived from the outer shell of sea or freshwater mollusks, is a special and important category of Chinese medicinal materials. To ensure the effective use of TTCM, a comparative identification study was performed on five commonly-used testacean drugs, including Haliotidis Concha, Arcae Concha, Meretricis Concha, Ostreae Concha and Margaritifera Concha(Shijueming, Walengzi, Geqiao, Muli and Zhenzhumu in Chinese, respectively).Typical morphological photographs of the crude drugs were acquired, and the key microscopic characteristics of the derived powders under normal light microscope and polarized light microscope were summarized. The major results can be concluded as follows:(1) the original species involved in the five TTCMs could be distinguished by their respective interspecies morphological characteristics;(2) the key identification characteristics of the five powdered crude drugs were mainly crystal fragments, with the fragment features under both normal light and polarized light microscope providing powerful points for differentiating the five commonly-used testacean drugs. This study demonstrated that it is feasible to provide authentication for these five kinds of TTCMs by the combination of morphology with microscopy.

胆石体外调节人体胆汁的胆固醇结晶途径

背景:胆固醇结晶是胆石形成的一个必要步骤。尽管一些模拟胆汁研究显示,胆石表面与周围水相对于结晶形成过程中的胆固醇分子存在竞争关系;但相关研究尚未在人体胆汁中开展。方法:收集13例行腹腔镜胆囊切除术患者的新鲜胆汁,其中10例为胆固醇结石,3例为胆色素结石。再从其他2例患者收集小的胆固醇结石。应用偏光显微镜观察原始胆汁和超滤胆汁(加入或不加入胆石)中胆固醇结晶的形态,包括螺旋状和针状无水胆固醇结晶,以及片状和块状胆固醇单水结晶。分别于胆石加入胆汁前和加入21天后,称量胆石的重量。结果:在原始胆汁中加入胆石后,无水胆固醇结晶有减少的趋势,但块状胆固醇单水结晶显著增多。在超滤胆汁中加入胆石后,螺旋状结晶和针状结晶的形成有受抑趋势,而片状和块状胆固醇单水结晶数量显著增多。胆汁孵育21天后,胆石重量显著减轻,无论是原始胆汁(-4.5%61.6%,P=0.046)还是超滤胆汁(-6.5%61.5%,P=0.002)。来自胆色素结石患者的胆汁,在未加入胆石前是清亮的;但加入胆固醇胆石后,所有样本均早期出现片状和块状胆固醇单水结晶。结论:无论是在原始胆汁还是超滤胆汁中,加入胆固醇结石均可对胆固醇结晶途径产生重大影响,胆固醇单水结晶的数量增加,而无水胆固醇结晶的形成受抑。固态胆固醇结晶从胆石表面分离或可解释这一现象。