Dark-field Light Microscopy Research Articles

ON FUNCTIONAL PURPOSE OF THE TALE SPIKE AND DORSAL SPINE IN PROTOSTRONGYLIDAE LARVAE

This article presents an attempt to analyze the functional purpose of the tail spike and dorsal spine of protostrongylid species. The actual material was obtained from the feces of reindeer (Rangifer tarandus) (the species Elaphostrongylus rangiferi was verified genetically), red deer (Cervus elaphus) and dapple deer (C. nippon) by the Vajda method and has been studied from 2018 to the present day by means of lightfield and dark-field light microscopy, and phase-contrast and scanning electron microscopy. It was found that the tail spike and dorsal spine 1) do not surpass the rest of the cuticle in their density (they are squashed by pressure of the cover slide, shriveled in preparation for scanning electron microscopy, and look uniform in relation to the body of the larva in polarized light); 2) have no openings in the apical areas. On this basis, the versions about the role of spines in episodes of intra- or interspecific competition and defensive function were rejected. The assumptions about the deposition (of substances or ultramicroscopic objects) and about fixation (linking) remain possible. The reliable purpose of the tail spike and dorsal spine of protostrongylid species larvae remains unknown. The further study of a connection between these larvae and their intermediate hosts, gastropods, seems promising.

In Operando Visualization of the Electrochemical Formation of Liquid Polybromide Microdroplets.

Zinc-bromine flow batteries are promising for stationary energy storage, and bromine-complexing agents have been used to form phase-separated liquid polybromide products. However, an understanding of the dynamics of polybromide nucleation is limited due to the beam sensitivity and complexity of polybromides. Here we report an in operando platform composed of dark-field light microscopy and a transparent electrochemical cell to reveal the dynamics of polybromide formation in their native environment. Using our platform, we confirm and reveal the liquid nature, chemical composition, pinning effect (strong interaction with Pt), residual effect (residual charge products on the surface), self-discharging, and over-oxidation of the polybromide products. The results provide insights into the role of complexing agents and guide the future design of zinc-bromine flow batteries. Furthermore, our in operando platform can potentially be used to study sensitive species and phases in other electrochemical reactions.

In Operando Optical Visualization of Br5 - electrochemistry with a Planar Glass Battery for Zn/Br Flow Batteries

Zn/Br flow batteries are one of the promising technologies for stationary energy storage. Bromine complexing agents have been used to form phase-separated liquid polybromide charging products. However, dynamic and microscopic understandings of the nucleation and growth of the polybromides are limiteddue to the beam-sensitivity and complexity of these compounds and their surrounding electrolytes. In this communication, we report an in operando platform composed of dark field light microscopy and a transparent planar electrochemical cell to dynamically reveal the electrochemical nucleation and formation of polybromides. Using our platform, we confirm the liquid nature, reveal the pinning effect (strong interaction with Pt), memory effect (surface residual), self-discharge of the polybromide product, and discover over-oxidation as a side reaction for ZBB. We also conclude that the electrochemically formed droplets are mainly MEPBr5,rather than previously reported MEPBr3 with in situ Raman spectroscopy. These results provide mechanistic insightsinto the role of ionic liquid complexing agents and Zn/Br battery design. The in operando visualization platform can also be applied to study other unstable or transient products or intermediates in electrochemical reactions in ambient environment.

Aquaporin 0 Modulates Lens Gap Junctions in the Presence of Lens-Specific Beaded Filament Proteins.

PurposeThe objective of this study was to understand the molecular and physiologic mechanisms behind the lens cataract differences in Aquaporin 0-knockout-Heterozygous (AQP0-Htz) mice developed in C57 and FVB (lacks beaded filaments [BFs]) strains.MethodsLens transparency was studied using dark field light microscopy. Water permeability (Pf) was measured in fiber cell membrane vesicles. Western blotting/immunostaining was performed to verify expression of BF proteins and connexins. Microelectrode-based intact lens intracellular impedance was measured to determine gap junction (GJ) coupling resistance. Lens intracellular hydrostatic pressure (HP) was determined using a microelectrode/manometer system.ResultsLens opacity and spherical aberration were more distinct in AQP0-Htz lenses from FVB than C57 strains. In either background, compared to wild type (WT), AQP0-Htz lenses showed decreased Pf (approximately 50%), which was restored by transgenic expression of AQP1 (TgAQP1/AQP0-Htz), but the opacities and differences between FVB and C57 persisted. Western blotting revealed no change in connexin expression levels. However, in C57 AQP0-Htz and TgAQP1/AQP0-Htz lenses, GJ coupling resistance decreased approximately 2.8-fold and the HP gradient decreased approximately 1.9-fold. Increased Pf in TgAQP1/AQP0-Htz did not alter GJ coupling resistance or HP.ConclusionsIn C57 AQP0-Htz lenses, GJ coupling resistance decreased. HP reduction was smaller than the coupling resistance reduction, a reflection of an increase in fluid circulation, which is one reason for the less severe cataract in C57 than FVB. Overall, our results suggest that AQP0 modulates GJs in the presence of BF proteins to maintain lens transparency and homeostasis.

Complementary Imaging of Silver Nanoparticle Interactions with Green Algae: Dark-Field Microscopy, Electron Microscopy, and Nanoscale Secondary Ion Mass Spectrometry.

Increasing consumer use of engineered nanomaterials has led to significantly increased efforts to understand their potential impact on the environment and living organisms. Currently, no individual technique can provide all the necessary information such as their size, distribution, and chemistry in complex biological systems. Consequently, there is a need to develop complementary instrumental imaging approaches that provide enhanced understanding of these "bio-nano" interactions to overcome the limitations of individual techniques. Here we used a multimodal imaging approach incorporating dark-field light microscopy, high-resolution electron microscopy, and nanoscale secondary ion mass spectrometry (NanoSIMS). The aim was to gain insight into the bio-nano interactions of surface-functionalized silver nanoparticles (Ag-NPs) with the green algae Raphidocelis subcapitata, by combining the fidelity, spatial resolution, and elemental identification offered by the three techniques, respectively. Each technique revealed that Ag-NPs interact with the green algae with a dependence on the size (10 nm vs 60 nm) and surface functionality (tannic acid vs branched polyethylenimine, bPEI) of the NPs. Dark-field light microscopy revealed the presence of strong light scatterers on the algal cell surface, and SEM imaging confirmed their nanoparticulate nature and localization at nanoscale resolution. NanoSIMS imaging confirmed their chemical identity as Ag, with the majority of signal concentrated at the cell surface. Furthermore, SEM and NanoSIMS provided evidence of 10 nm bPEI Ag-NP internalization at higher concentrations (40 μg/L), correlating with the highest toxicity observed from these NPs. This multimodal approach thus demonstrated an effective approach to complement dose-response studies in nano-(eco)-toxicological investigations.

Alkaliphilic Bacillus species show potential application in concrete crack repair by virtue of rapid spore production and germination then extracellular calcite formation.

Characterization of alkaliphilic Bacillus species for spore production and germination and calcite formation as a prelude to investigate their potential in microcrack remediation in concrete. Conditions, extent and timing of endospore production was determined by dark-field light microscopy; germination induction and kinetics were assessed by combining reduction in optical density with formation of refractile bodies by phase-contrast microscopy. Bacillus pseudofirmus was selected from several species as the most suitable isolate. Levels and timing of calcium carbonate precipitated invitro by B. pseudofirmus were evaluated by atomic absorption spectroscopy and structural identity confirmed as calcite and aragonite by Raman spectroscopy and FTIR. The isolate produced copious spores that germinated rapidly in the presence of germinants l-alanine, inosine and NaCl. Bacterial cells produced CaCO3 crystals in microcracks and the resulting occlusion markedly restricted water ingress. By virtue of rapid spore production and germination, calcium carbonate formation invitro and insitu, leading to sealing of microcracks, B. pseudofirmus shows clear potential for remediation of concrete on a commercial scale. Microbial sealing of microcracks should become a practicable and sustainable means of increasing concrete durability.

Method to extract minimally damaged collagen fibrils from tendon.

A new method is presented to extract collagen fibrils from mammalian tendon tissue. Mammalian tendons are treated with a trypsin-based extraction medium and gently separated with tweezers in an aqueous solution. Collagen fibrils released in the solution are imaged using both dark-field light microscopy and scanning electron microscopy. The method successfully extracts isolated fibrils from rat tail and patellar tendons. To examine whether the method is likely to damage fibrils during extraction, sea cucumber dermis fibril lengths are compared against those obtained using only distilled water. The two methods produce fibrils of similar lengths. This is contrasted with fibrils being shortened when extracted using a tissue homogenizer. Scanning electron microscopy shows the new method preserves D-banding features on fibril surfaces and that fibril diameter does not vary substantially compared with water extracted fibrils.

Silver Nanoparticle-Induced Autophagic-Lysosomal Disruption and NLRP3-Inflammasome Activation in HepG2 Cells Is Size-Dependent.

Silver nanoparticles (AgNPs) are incorporated into medical and consumer products to exploit their excellent antimicrobial properties; however, potential mechanisms of toxicity of AgNPs in mammalian cells are not fully understood. The objective of this study was to determine the mechanism of size- and concentration-dependent cytotoxicity of AgNPs in human liver-derived hepatoma (HepG2) cells. Mechanisms of toxicity were explored at subcytotoxic concentrations (≤10 µg/ml AgNPs) and autophagy induction, lysosomal activity, inflammasome-dependent caspase-1 activation, and apoptosis were examined. Using enhanced dark-field light microscopy, hyperspectral imaging, electron microscopy, and energy dispersive X-ray spectroscopy, AgNPs were shown to rapidly accumulate in cytoplasmic vesicles for up to 24 h and 10-nm AgNPs exhibited the highest uptake and accumulation. Autophagy and enhanced lysosomal activity were induced at noncytotoxic concentrations (1 µg/ml; primary particle size:10 > 50 >100 nm), whereas increased caspase-3 activity (associated with apoptosis) was observed at cytotoxic concentrations (10, 25, and 50 µg/ml). Subcytotoxic concentrations of AgNPs enhanced expression of LC3B, a pro-autophagic protein, and CHOP, an apoptosis inducing ER-stress protein, and activation of NLRP3-inflammasome (caspase-1, IL-1β). Disrupting the autophagy-lysosomal pathway through chloroquine or ATG5-siRNA exacerbated AgNPs-induced caspase-1 activation and lactate dehydrogenase release, suggesting that NLRP3-inflammasome plays an important role in AgNPs-induced cytotoxicity. Overall, 10-nm AgNPs showed the highest cellular responses compared with 50- and 100-nm AgNPs based on equal mass dosimetry. The results indicate the potential of vesicle-engulfed 10-nm AgNPs to induce cytotoxicity by a mechanism involving perturbations in the autophagy-lysosomal system and inflammasome activation.

Charcoal recognition, taphonomy and uses in palaeoenvironmental analysis

Charcoal, predominantly the product of wildfires, is abundant in many sedimentary rocks deposited in a wide range of environments, from terrestrial to marine. It also occurs in some volcanic rocks. This paper outlines aspects of charcoal formation (both natural and experimental) and briefly considers the taphonomic processes leading to a final assemblage. This is done using examples from recent fires and through experimentation. The ways in which charcoal assemblages are recognized in the field and extraction in the laboratory are also considered. Methods of charcoal identification are presented. The range of charcoalified plant organs that can be found is discussed and a wide range of study methods outlined (including light microscopy, dark field light microscopy, reflectance microscopy, scanning electron microscopy and synchrotron radiation X-ray tomographic microscopy). Emphasis in this paper is on the study of macro- and meso-charcoal (above180 µm). Finally there is a consideration of the broad use of charcoal from plant evolution studies, fire history studies, vegetation studies, anatomical studies, climate and atmospheric studies and the wider importance of charcoal for the Earth and Biological Sciences. Charcoal is information rich but yet is an under-utilized resource.

10.1007/s11449-008-2023-8

The collective plasmon resonances in a monolayer formed by metal or metal-dielectric nanoparticles with dipole or quadrupole single-particle resonances are theoretically and experimentally studied. The extinction, scattering, and absorption spectra are calculated using an exact many-particle solution for the system of interacting particles. With increasing surface density of particles in the monolayer, the dipole resonance is suppressed, and the spectrum of the collective system is determined by the quadrupole plasmon only. It is shown that the selective suppression of the long-wavelength extinction band is caused by the collective suppression of the dipole scattering mode, whereas the short-wavelength absorption spectrum of the monolayer differs little from the single-particle spectrum. Using dark-field light and atomic force microscopy, the kinetics of self-assembling of nanoshells is studied. It is shown that the universal linear relation between the relative shift of the wavelength of the collective quadrupole resonance and the relative increment of the refractive index of the surrounding medium is implemented.

Collective plasmon resonances in monolayers of metal nanoparticles and nanoshells

The collective plasmon resonances in a monolayer formed by metal or metal-dielectric nanoparticles with dipole or quadrupole single-particle resonances are theoretically and experimentally studied. The extinction, scattering, and absorption spectra are calculated using an exact many-particle solution for the system of interacting particles. With increasing surface density of particles in the monolayer, the dipole resonance is suppressed, and the spectrum of the collective system is determined by the quadrupole plasmon only. It is shown that the selective suppression of the long-wavelength extinction band is caused by the collective suppression of the dipole scattering mode, whereas the short-wavelength absorption spectrum of the monolayer differs little from the single-particle spectrum. Using dark-field light and atomic force microscopy, the kinetics of self-assembling of nanoshells is studied. It is shown that the universal linear relation between the relative shift of the wavelength of the collective quadrupole resonance and the relative increment of the refractive index of the surrounding medium is implemented.

Luminance Contrast — a New Visible Light Technique for Examining Transparent Specimens

Abstract Transparent specimens are usually examined by dark field, phase contrast and interference contrast light microscopy. In dark field, specimens are illuminated by oblique light beams that come from the periphery of the illuminating apparatus. Therefore, some transparent objects, e.g. unstained native bacteria, are barely visible and fine structures inside them are often not visible. In phase contrast, the discernment of fine detail can be reduced by halo artifacts. The intensity of contrast, i.e. the difference in brightness between the background and specimen, is constant and not variable; it is determined by the specification of the phase ring within the phase contrast lens and dependent on the specific phase differences between the specimen and its surrounding medium. Interference contrast images are free from halo artifacts, but their contrast may be lower than in corresponding phase contrast or dark field images, especially, when transparent specimens are examined in thinlayer preparations.

Modifications of stereological correction methods for precipitate parameters using transmission microscopy

Energy filtered transmission electron microscopy (EFTEM), dark field imaging and light microscopy are well established methods to evaluate size, volume fraction and number density of precipitates on different size scales. The stereological problems are well known and analogous for all methods. Many evaluation methods can be found in literature, each of them valid for special sample geometries or special size distributions. Despite this variety, serious errors can occur if the assumptions do not exactly represent the real situation. In this paper, available and newly developed correction methods are tested for evaluating these parameters for different precipitate size distributions in a sample of arbitrary topography. It is shown that the newly proposed methods produce significantly lower deviations and are more assessable regarding their systematic errors.

Dopamine D1/5 Receptor Modulation of Firing Rate and Bidirectional Theta Burst Firing in Medial Septal/Vertical Limb of Diagonal Band Neurons In Vivo

The medial septum/vertical limb of diagonal band complex (MS/vDB) consists of cholinergic, GABAergic, and glutamatergic neurons that project to the hippocampus and functionally regulate attention, memory, and cognitive processes. Using tyrosine hydroxlase (TH) immunocytochemistry and dark-field light microscopy, we found that the MS/vDB is innervated by a sparse network of TH-immunoreactive (putative catecholaminergic) terminals. MS/vDB neurons are known to fire in rhythmic theta burst frequency of 3-7 Hz to pace hippocampal theta rhythm. Extracellular single-unit recording in theta and non-theta firing MS/vDB neurons and antidromically identified MS/vDB-hippocampal neurons were made in urethan-anesthetized rats. Tail-pinch noxious stimuli and ventral tegmental area (VTA) stimulation (20 Hz) evoked spontaneous theta burst firing in MS/vDB neurons. Systemic D1/5 antagonists SCH23390 or SCH39166 (0.1 mg/kg iv) alone suppressed the spontaneous theta bursts, suggesting a tonic facilitatory endogenous dopamine D1 "tone" that modulates theta bursts in vivo. Activation of D1/5 receptor by dihydrexidine (10 mg/kg iv) led to an increase in mean firing rate in 60% of all theta and non-theta MS/vDB neurons with an increase in the number of theta bursts and spikes/burst in theta cells. In strong theta firing MS/vDB neurons, D1/5 receptor stimulation suppressed the occurrence of theta burst firing, whereas the overall increase in spontaneous mean firing rate remained. In low baseline theta MS/vDB neurons D1/5 receptor stimulation increases the occurrence of theta bursts along with a net increase in mean firing rate. Atropine injection consistently disrupts theta burst pattern and reduced the time spent in theta firing. Collectively, these data suggest that dopamine D1/5 stimulation enhances the mean firing rate of most MS/vDB neurons and also provides a state-dependent bidirectional modulation of theta burst occurrence. Some of these MS/vDB neurons may be cholinergic or GABAergic that may indirectly regulate theta rhythm in the hippocampus.

Mechanical analyses of morphological and topological transformation of liposomes

Liposomes are micro-compartments made of lipid bilayer membranes possessing the characteristics quite similar to those of biological membranes. To form artificial cell-like structures, we made liposomes that contained subunit proteins of cytoskeletons: tubulin or actin. Spherical liposomes were transformed into bipolar or cell-like shapes by mechanical forces generated by the polymerization of encapsulated subunits of microtubules. On the other hand, disk- or dumbbell-shaped liposomes were developed by the polymerization of encapsulated actin. Dynamic processes of morphological transformations of liposomes were visualized by high intensity dark-field light microscopy. Topological changes, such as fusion and division of membrane vesicles, play an essential role in cellular activities. To investigate the mechanism of these processes, we visualized the liposomes undergoing topological transformation in real time. A variety of novel topological transformations were found, including the opening-up of liposomes and the direct expulsion of inner vesicles.

The structure of the helically perturbed flagellar filament of Pseudomonas rhodos: implications for the absence of the outer domain in other complex flagellins and for the flexibility of the radial spokes.

Bacterial flagella, the organelles of motility, are commonly divided into two classes: 'plain' and 'complex'. The complex filaments are pairwise, helically perturbed forms of the plain filaments and have been reported to occur only in Rhizobium and Pseudomonas. Previously, we reconstructed and analysed the structure of the complex filaments of Rhizobium lupini H13-3 and determined their unique symmetry and origin of the perturbations (Trachtenberg et al., 1986, J Mol Biol 190: 569-576; 1987, 195: 603-620; 1998, 276: 759-773; Cohen-Krausz and Trachtenberg, 1998, J Struct Biol 122: 267-282). Here, we analyse the structure of the flagellar filament of the other known complex filament, that of Pseudomonas rhodos, as reconstructed from electron microscope images. Compared with the filament of R. lupini, the filament of P. rhodos is more flexible, as implied from high-intensity darkfield light microscopy and, although constructed from flagellins of higher molecular weights (59 versus 41 kDa), has similar symmetry. Using cryonegative stained specimens and low-dose, field emission electron microscopy, we reconstructed and averaged 158 filaments each containing 170 statistically significant layer lines. The three-dimensional density maps of P. rhodos clearly suggest, when compared with those of R. lupini and the right-handed Salmonella typhimurium SJW1655, that R. lupini is missing the outer flagellin domain (D3), that the interior of the complex filament is rather similar to that of the plain filament and that the radial spokes (connecting domains D0 and D1), present in individual density maps, average out because of their variability and implied flexibility. Extending the three-start grooves and ridges on the propeller's surface, in the form of an Archimedean screw, may further improve the motility of the cell in viscous environments.

The bacterial linear motor of Spiroplasma melliferum BC3: from single molecules to swimming cells.

Spiroplasma melliferum BC3 are wall-less bacteria with internal cytoskeletons. Spiroplasma, Mycoplasma and Acholeplasma belong to the Mollicutes, which represent the smallest, simplest and minimal free-living and self-replicating forms of life. The Mollicutes are motile and chemotactic. Spiroplasma cells are, in addition, helical in shape. Based on data merging, obtained by video dark-field light microscopy of live, swimming helical Spiroplasma cells and by cryoelectron microscopy, unravelling the subcellular structure and molecular organization of the cytoskeleton, we propose a functional model in which the cytoskeleton also acts as a bacterial linear motor enabling and controlling both dynamic helicity and swimming. The cytoskeleton is a flat, monolayered ribbon constructed from seven contractile fibrils (generators) capable of changing their length differentially in a co-ordinated manner. The individual, flat, paired fibrils can be viewed as chains of tetramers approximately 100 A in diameter composed of 59 kDa monomers. The cytoskeletal ribbon is attached to the inner surface of the cell membrane (but is not an integral part of it) and follows the shortest helical line on the coiled cellular tube. We show that Spiroplasma cells can be regarded, at least in some states, as near-perfect dynamic helical tubes. Thus, the analysis of experimental data is reduced to a geometrical problem. The proposed model is based on simple structural elements and functional assumptions: rigid circular rings are threaded on a flexible, helical centreline. The rings maintain their circularity and normality to the centreline at all helical states. An array of peripheral, equidistant axial lines forms a regular cylindrical grid (membrane), by crossing the lines bounding the rings. The axial and peripheral spacing correspond to the tetramer diameter and fibril width (100 A) respectively. Based on electron microscopy data, we assign seven of the axial grid lines in the model to function as contractile generators. The generators are clustered along the shortest helical paths on the cellular coil. In the model, the shortest generator coincides with the shortest helical line. The rest, progressively longer, six generators follow to the right or to the left of the shortest generator in order to generate the maximal range of lengths. A rubbery membrane is stretched over (or represented by) the three-dimensional grid to form a continuous tube. Co-ordinated, differential length changes of the generators induce the membranal cylinder to coil and uncoil reversibly. The switch of helical sense requires equalization of the generators' length, forming a straight cylindrical tube with straight generators. The helical parameters of the cell population, obtained by light microscopy, constitute several subpopulations related, most probably, to cell size and age. The range of molecular dimensions in the active cytoskeleton inferred from light microscopy and modelling agrees with data obtained by direct measurements of subunit images on electron micrographs, scanning transmission electron microscopy (STEM) and diffraction analysis of isolated ribbons. Swimming motility and chemotactic responses are carried out by one or a combination of the following: (i). reciprocating helical extension and compression ('breathing'); (ii). propagation of a deformation (kink) along the helical path; (iii). propagation of a reversal of the helical sense along the cell body; and (iv). irregular flexing and twitching, which is functionally equivalent to standard bacterial tumbling. Here, we analyse in detail only the first case (from which all the rest are derived), including switching of the helical sense.

On Visualization of Sub-Micron Particles with Dark-Field Light Microscopy

Dark-field light microscopy is widely employed to visualize colloidal particles much smaller than the light wavelength used. In the captured images, the colloidal particles appear, against a dark background, as bright “specks” much larger than the geometrical size of the particles. To verify whether the “specks” are for individual particles or clusters of particles, experiments are performed which used low bulk concentrations of five suspensions of monodispersed particles (∼0.3μm in diameter) and a dark-field video microscopic system with an optical resolution of ∼0.5μm to count the particles after they all have deposited onto the inner surfaces of a parallel-plate glass channel. The average size and the size distribution of the particles are also determined at the end of each experiment. The results confirmed that the visualized “specks” are for individual particles. The measured and prepared particle bulk concentrations in the five experiments closely matched, to within ±5%, and the measured average size of the particles and their size distribution at the end of the five experiments were in agreement with the known values.

Automated Video Microscopic Imaging and Data Acquisition System for Colloid Deposition Measurements

An optical video microscopic system and image processing and data extraction and manipulation routines are developed for in situ detailed quantification of the deposition of colloids onto an arbitrary surface and determining their concentration distribution across the bulk suspension. The system produces a relatively large field of view (∼330×245μm) and utilizes dark-field light microscopy to visualize colloids as small as ∼0.3μm in diameter at the surface and in the bulk suspension with a sufficient resolution (∼0.5μm). On real-time basis, the routines automate various tasks from image capturing and processing to data manipulation, extraction, and display. The extracted data include: (i) surface concentration and flux of deposited, attached, and detached colloids, (ii) surface concentration and flux of arriving and departing colloids, (iii) distribution of colloids in the bulk suspension in the direction perpendicular to the deposition surface, and (iv) spatial and temporal distributions of deposited colloids. This article provides detailed description of the system and its image processing and data extraction and manipulation routines. Representative results of the deposition of 0.3-μm-diameter polystyrene colloids onto a glass surface, from a flowing suspension in a 0.02-cm-aperture parallel-plate channel, are presented and discussed.

LARGE DEFORMATION OF SPHERICAL VESICLE STUDIED BY PERTURBATION THEORY AND SURFACE EVOLVER

With tangent angle perturbation approach the axial symmetry deformation of a spherical vesicle in large under the pressure changes is studied by the elasticity theory of Helfrich spontaneous curvature model. Three main results in axial symmetry shape: biconcave shape, peanut shape, and one type of myelin are obtained. These axial symmetry morphology deformations are in agreement with those observed in lipsome experiments by dark-field light microscopy and in the red blood cell with two thin filaments (myelin) observed in living state. Furthermore, the biconcave shape and peanut shape can be simulated with the help of a powerful software, Surface Evolver, in which the spontaneous curvature can be easy taken into account.