Pore Cap Research Articles

Hollow polypyrrole coated with mesoporous silica nanoparticles graft copolymer multifunctional nanocomposites for intracellular cancer therapy

A novel hollow polypyrrole coated by mesoporous silica nanoparticles graft poly(2-(4-formylbenzoyloxy)ethyl methacrylate- co -2-(dimethylamino)ethyl methacrylate)- block -poly(triethylene glycol methyl ether methacrylate) multifunctional nanocomposite (HPPy@MSN-SS-NH- g -P(FBEMA- co -DMAEMA)- b -PTEGMA) was designed and prepared via disulfide bond linkages. This material as a gatekeeper integrated multifunctionality such as pore capping, and pH and potothermal multistimuli response abilities, and possessed good photothermal stability, near-infrared (NIR) photothermal conversion capacity and dispersion stability in aqueous solution. It could entrap anticancer doxorubicin (DOX) by pore channels of HPPy@MSN, the core of amphiphilic copolymer micelles and Schiff base linkages, offering a load capacity and load efficiency of up to about 41.5 and 83.1%, respectively. Dynamic light scattering analysis showed that the DOX-loaded preparations had particle size of less than 300 nm, and could achieve intracellular chemo–photothermal combination therapy by controlling the mesopore on-off based on the gatekeepers, tumor environments including pH and reductant stimuli. The drug-loaded nanoparticles exhibited optimal drug release dynamics in cancer tissues upon triggered by pH and glutathione (GSH) under the NIR irradiation. Cell counting Kit-8 assay manifested that the nanocarriers are non-toxic and safe, whereas the drug-loaded preparations exhibited robust anticancer efficacy. Cellular uptake and TUNEL assay revealed high concentrations of DOX accumulating around and inside Hela cells, accordingly producing significant tumor cell apoptosis. Therefore, the newly-developed materials can be used for comprehensive therapy of cancer as a promising candidate of drug delivered carriers. • We prepared hollow PPy coated with MSNs graft copolymer multifunctional composites. • The nanocomposites exhibited pH, reductant and potothermal multi-responsivities. • This material entrapped DOX by pores, hydrophobic actions and Schiff base linkages. • The drug release was triggered by synergism of pore capping, pH, reductant and NIR. • A chemo–photothermal combination therapy could be achieved by the mesopore on-off.

PENDEKATAN VARIABEL COST DALAM RANCANGAN SISTEM INFORMASI BIAYA PRODUKSI BUSANA MUSLIM

One method in determining the cost of goods is the variable costing method. This method determining the cost of goods manufactured which classifies costs based on cost behavior consisting of raw material costs, direct labor costs, and factory overhead costs. By using the variable costing method, fixed costs are separated into periodic costs which are used as a deduction for the contribution margin in the profit/loss statement. The contribution margin is the remaining amount from sales after deducting variable costs. This amount contributes to covering fixed costs and generating a profit for a certain period. If the company can calculate the cost of goods manufactured correctly and precisely, it will certainly assist management in carrying out the process of planning, controlling, or controlling production costs, determining the right selling price, and planning a good profit. This study builds a Muslim clothing production cost information system using the variable cost method. The new design of information system consists of 5 input data (Model, Material Cost, Consumer, Payment, PO), seventh output data (Consumer Report, Payment Report, Receipt, List of Clothing Models, PO Recap, Production Report, Production Cost Report) and single database with five tables (PO, Payment, Consumer, Product, Stock).

Continuous polymer films deposited on top of porous substrates using plasma-enhanced atomic layer deposition and molecular layer deposition

Molecular layer deposition (MLD) is able to produce ultrathin polymer films with control over thickness, cross-linking, and chemical composition. With these capabilities, MLD should be useful in the fabrication of novel polymer membranes on porous supports. However, confining a continuous MLD film to the surface of porous substrates is difficult because of MLD film growth in the pores. The deposition in the pores lowers the conductance of the porous support. This paper presents a method to deposit continuous polymer films on top of porous substrates. In this method, Al2O3 plasma-enhanced atomic layer deposition (PE-ALD) using trimethylaluminum and oxygen plasma as the reactants was first used to cap the pores of the substrate. Subsequently, a polyamide MLD film was deposited on the Al2O3 PE-ALD capping layer using m-phenylenediamine and trimesoyl chloride as the reactants. The Al2O3 pore caps were then removed from the porous substrate by etching from the backside using a timed exposure to a dilute sodium hydroxide solution. This method was demonstrated using anodic aluminum oxide (AAO) and polyethersulfone (PES) porous substrates. Al2O3 PE-ALD film growth was limited to the top of the porous substrate, resulting in rapid surface recombination or high sticking coefficients for the reactive plasma species within the pores. Gas permeance measurements confirmed the pore capping of the AAO substrates. The reopening of the pores by dissolving the Al2O3 pore caps with a sodium hydroxide solution was monitored using gas permeance versus etch time. The removal of the Al2O3 pore caps from the PES substrates could also dissolve the Al2O3 layer underneath the MLD film. The loss of this Al2O3 layer led to the detachment of the MLD film from the PES substrate. However, the MLD film could be anchored to the PES support at fractures located in the Al2O3 film prior to the MLD. The Al2O3 film fracture allowed the MLD film to anchor firmly to the PES substrate by MLD in the pores of the PES porous substrate. The distance between the anchor points was a function of fracture density. This distance could be controlled by applying a tensile stress to the Al2O3 PE-ALD film to fracture the film through sample bending. This method produced firmly anchored polymer MLD films on top of the PES porous substrates.

Supramolecular Nanomachines as Stimuli-Responsive Gatekeepers on Mesoporous Silica Nanoparticles for Antibiotic and Cancer Drug Delivery.

Major objectives in nanomedicine and nanotherapy include the ability to trap therapeutic molecules inside of nano-carriers, carry therapeutics to the site of the disease with no leakage, release high local concentrations of drug, release only on demand - either autonomous or external, and kill the cancer cells or an infectious organism. This review will focus on mesoporous silica nanoparticle carriers (MSN) with a large internal pore volume suitable for carrying anticancer and antibiotic drugs, and supramolecular components that function as caps that can both trap and release the drugs on-command.Caps that are especially relevant to this review are rotaxanes and pseudorotaxanes that consist of a long chain-like molecule threaded through a cyclic molecule. Under certain conditions discussed throughout this review, the cyclic molecule can be attracted to one end of the rotaxane and in the presence of a stimulus can slide to the other end. When the thread is attached near the pore opening on MSNs, the sliding cyclic molecule can block the pore when it is near the particle or open it when it slides away.The design, synthesis and operation of supramolecular systems that act as stimuli-responsive pore capping devices that trap and release molecules for therapeutic or imaging applications are discussed. Uncapping can either be irreversible because the cap comes off, or reversible when the cyclic molecule is prevented from sliding off by a steric barrier. In the latter case the amount of cargo released (the dose) can be controlled. These nanomachines act as valves.Examples of supramolecular systems stimulated by chemical signals (pH, redox, enzymes, antibodies) or by external physical signals (light, heat, magnetism, ultrasound) are presented. Many of the systems have been studied in vitro proving that they are taken up by cancer cells and release drugs and kill the cells when stimulated. Some have been studied in mouse models; after IV injection they shrink tumors or kill intracellular pathogens after stimulation. Supramolecular constructs offer fascinating, highly controllable and biologically compatible platforms for drug delivery.

Hyaluronic acid coated hollow mesoporous silica nanoparticles for reduction‐triggered drug release

In this work, the reduction-responsive hyaluronic acid (HA) coated hollow mesoporous silica nanoparticles (HMSNs) were developed for the targeted tumour therapy. First, the anticancer drug doxorubicin was retained inside the central hollow cavities of the HMSNs with high loading efficiency. Meanwhile, the adamantane group was anchored on the surface of the HMSNs using the disulphide bond as the reduction–cleavable linkage. Subsequently, the β-cyclodextrin (β-CD) decorated HA was immobilised on the surface of HMSNs by the strong β-CD/adamantane host–guest interactions. Specifically, the β-CD acted as the pore caps to block the loaded drug inside, while HA served as the targeting ligand for the effective cancer therapy. Under the intracellular reductive environment, the nanocomposites displayed reduction-triggered drug release manner. Confocal microscopy observations confirmed the nanocomposites were easily internalised by HeLa cancer cells and released the loaded drug inside. In addition, the cytotoxicity study evidenced that HA decorated nanocomposites exhibited improved biocompatibility and antitumour activity as compared to that without HA modification. Thus, the reduction-responsive nanocomposites could be promising as a drug delivery platform toward the cancer therapy.

Septal-pore-associated structures of Hysterangium clathroides and Hysterangium nephriticum (Hysterangiales, Basidiomycota, Fungi)

The hyphae of Basidiomycota and Ascomycota are divided into compartments by the cross-walls (septa). In mushroom-forming fungi, members of subphylla Pezizomycotina O.E. Erikss. & Winka (Ascomycota) and Agaricomycotina Doweld (Basidiomycota), the septa are associated with specific intracellular structures, which play a role in maintaining the integrity of hyphae in case of injury but also in cell differentiation (Webster & Weber 2007, Moore et al. 2011). In basidiomycetes, these structures may also be involved in certain intracellular processes and the formation of sporocarps (Van Peer et al. 2010, Moore et al. 2011). In this fungal group they are the reticulum-connected and -derived membranous structures called septal pore caps (SPC) or parenthesomes, associated with a dolipore (Moore 1985, 1996, van Driel 2007). The pores may also be occluded by a range of materials of different form. The septal-pore-associated structures may differ considerably between species of different evolutionary lineages. This quality was used as a differentiating factor in the taxonomy at both species and higher-order levels (Lutzoni et al. 2004, McLaughlin et al. 2009).

Extremely thin layer plastification for focused-ion beam scanning electron microscopy: an improved method to study cell surfaces and organelles of cultured cells.

Since the recent boost in the usage of electron microscopy in life-science research, there is a great need for new methods. Recently minimal resin embedding methods have been successfully introduced in the sample preparation for focused-ion beam scanning electron microscopy (FIB-SEM). In these methods several possibilities are given to remove as much resin as possible from the surface of cultured cells or multicellular organisms. Here we introduce an alternative way in the minimal resin embedding method to remove excess of resin from two widely different cell types by the use of Mascotte filter paper. Our goal in correlative light and electron microscopic studies of immunogold-labelled breast cancer SKBR3 cells was to visualise gold-labelled HER2 plasma membrane proteins as well as the intracellular structures of flat and round cells. We found a significant difference (p < 0.001) in the number of gold particles of selected cells per 0.6 μm2 cell surface: on average a flat cell contained 2.46 ± 1.98 gold particles, and a round cell 5.66 ± 2.92 gold particles. Moreover, there was a clear difference in the subcellular organisation of these two cells. The round SKBR3 cell contained many organelles, such as mitochondria, Golgi and endoplasmic reticulum, when compared with flat SKBR3 cells. Our next goal was to visualise crosswall associated organelles, septal pore caps, of Rhizoctonia solani fungal cells by the combined use of a heavy metal staining and our extremely thin layer plastification (ETLP) method. At low magnifications this resulted into easily finding septa which appeared as bright crosswalls in the back-scattered electron mode in the scanning electron microscope. Then, a septum was selected for FIB-SEM. Cross-sectioned views clearly revealed the perforate septal pore cap of R. solani next to other structures, such as mitochondria, endoplasmic reticulum, lipid bodies, dolipore septum, and the pore channel. As the ETLP method was applied on two widely different cell types, the use of the ETLP method will be beneficial to correlative studies of other cell model systems and multicellular organisms.

Effects of Bubble Location on Pore Shape in Solid

AbstractThe shapes of a pore resulting from an entrapped bubble by a solidification front for different locations of the bubble below the free surface are predicted in this work. Bubble location is an important factor affecting temperature gradient in liquid, solute gas dissipated into the ambient, heterogeneous nucleation of the bubble and shape of the bubble cap, and subsequent entrapment and the pore shape in solid. The shapes of pores in solid influence not only material properties, but also contemporary issues of engineering, biology, medical technology and science, etc. This study takes into account solute transport across a coupling shape of the pore cap determined by the Young-Laplace equation governing balance of liquid, gas and capillary pressures. The results find that increases in depthwise location of a bubble increase pore radius and time for bubble entrapment as solute transport is from the pore across cap emerged through a concentration boundary layer along the solidification front into surrounding liquid in the early stage. On the other hand, the bubble cannot be entrapped, provided that solute transport in opposite directions across the cap submerged in a concentration boundary layer along the solidification front. The predicted growth and entrapment of a tiny bubble as a pore in solid agree with experimental data. Understanding and controlling of the pore shape via controlling bubble location is of interest and challenging.

Bond number effects on pore shape in solid

The pore shape affected by Bond number, indicating the ratio between hydrostatic pressure and capillary pressure, of a bubble entrapped by a solidification front is predicted. Bond number is responsible for bubble nucleation, surface area and shape of the bubble or pore cap beyond the solidification front. Surface area determines solute transport across the cap, whereas contact angle of the cap delineates pore shape in solid. The critical velocity for entrapment of a bubble is also related to Bond number. Pore formation in solid influence not only microstructure of materials, but also contemporary issues of biology, engineering, foods, geophysics and climate change, etc. This work extends previous models accounting for mass and momentum transport of solute across a coupled shape of the bubble cap to predict the pore shape in solid. The study also accounts for different directions and magnitudes of solute transport across the cap in different cases. Case 1 is subject to solute transport from the pore across an emerged cap in a concentration boundary layer on the solidification front to surrounding liquid in the early stage. Case 2 indicates solute transport across a submerged cap in a concentration boundary layer to the pore. In contrast to Case 2a, pore value expansion in Case 2b exhibits a more important role in solute concentration at the cap than solute transport across the cap in the late stage. The results show that an increase in Bond number decreases pore radius and time for bubble entrapment in Case 1. An isolated pore cannot be formed in Cases 2a and 2b. The predicted growth and entrapment of a tiny bubble as a pore in solid are found to agree with experimental data. A realistic prediction and control of the pore shape in solid via Bond number has therefore been obtained.

Biodegradable Oxamide-Phenylene-Based Mesoporous Organosilica Nanoparticles with Unprecedented Drug Payloads for Delivery in Cells.

We describe biodegradable mesoporous hybrid nanoparticles (NPs) in the presence of proteins and their applications for drug delivery. We synthesized oxamide phenylene-based mesoporous organosilica nanoparticles (MON) in the absence of a silica source which had remarkably high organic content and high surface areas. Oxamide functions provided biodegradability in the presence of trypsin model proteins. MON displayed exceptionally high payloads of hydrophilic and hydrophobic drugs (up to 84 wt %), and a unique zero premature leakage without the pore capping, unlike mesoporous silica. MON were biocompatible and internalized into cancer cells for drug delivery.

Co-loading of coralyne and indocyanine green into adenine DNA-functionalized mesoporous silica nanoparticles for pH- and near-infrared-responsive chemothermal treatment of cancer cells.

Despite remarkable progress in the construction of functional mesoporous silica nanoparticles (MSNs) for cancer therapy, a multifunctional system with synergistic advantages over any single model remains encouraging. The objective of the present study was to develop a novel, simple and powerful nanotherapeutic system for the pH- and near-infrared (NIR)-responsive chemothermal treatment of cancer cells by co-loading coralyne and indocyanine green (ICG) into the adenine DNA (poly(A))-functionalized MSN. Coralyne, a type of planar alkaloid for cancer chemotherapy, can not only be loaded into the pores of MSN, but can also bind poly(A) and trigger the formation of non-Watson-Crick secondary structures, resulting in pore capping by the cooperative binding of poly(A) strands. Under low-pH or high-temperature conditions, non-Watson-Crick secondary structures were unstable, leading to the dehybridization of the cooperative binding structure and open-gate state. ICG, another cargo co-loaded into MSN, showed high efficiency for the conversion of NIR light into heat, which could provide the fundamental basis of hyperthermal therapy and promote pore opening upon NIR irradiation. In vitro studies using human hepatoma (HepG-2) cells demonstrated that this system could perform well in cellular acidic environment and under NIR irradiation, leading to a significant efficiency in controlled drug release and chemothermal cancer treatment. We believe that with these excellent features it should become a favorable nanoplatform for biomedical applications.

Three dimensional structure of the anthrax toxin translocon–lethal factor complex by cryo‐electron microscopy

We have visualized by cryo-electron microscopy (cryo-EM) the complex of the anthrax protective antigen (PA) translocon and the N-terminal domain of anthrax lethal factor (LF(N) inserted into a nanodisc model lipid bilayer. We have determined the structure of this complex at a nominal resolution of 16 Å by single-particle analysis and three-dimensional reconstruction. Consistent with our previous analysis of negatively stained unliganded PA, the translocon comprises a globular structure (cap) separated from the nanodisc bilayer by a narrow stalk that terminates in a transmembrane channel (incompletely distinguished in this reconstruction). The globular cap is larger than the unliganded PA pore, probably due to distortions introduced in the previous negatively stained structures. The cap exhibits larger, more distinct radial protrusions, previously identified with PA domain three, fitted by elements of the NMFF PA prepore crystal structure. The presence of LF(N), though not distinguished due to the seven-fold averaging used in the reconstruction, contributes to the distinct protrusions on the cap rim volume distal to the membrane. Furthermore, the lumen of the cap region is less resolved than the unliganded negatively stained PA, due to the low contrast obtained in our images of this specimen. Presence of the LF(N) extended helix and N terminal unstructured regions may also contribute to this additional internal density within the interior of the cap. Initial NMFF fitting of the cryoEM-defined PA pore cap region positions the Phe clamp region of the PA pore translocon directly above an internal vestibule, consistent with its role in toxin translocation.

Fungal evo–devo: organelles and multicellular complexity

Peroxisome-derived Woronin bodies of the Ascomycota phyla, and the endoplasmic reticulum (ER)-derived septal pore cap (SPC) of the Basidiomycota, are both fungal organelles that prevent cytoplasmic bleeding when multicellular hyphal filaments are wounded. Analysis of Woronin body constituent proteins suggests that these organelles evolved in part through gene duplication and co-opting of non-essential genes for new functions, indicating that new organelles can arise through typical evolutionary mechanisms. Interestingly, clades possessing the Woronin body and SPC also produce the largest and most complex multicellular fungal reproductive structures. Certain Woronin body and SPC mutants have defects in growth and development, suggesting functions beyond cellular wound healing. I argue that studying these specialized systems will help to reveal the basis for fungal diversity and provide general principles for co-evolution of organelles and multicellular complexity.

The septal pore cap is an organelle that functions in vegetative growth and mushroom formation of the wood‐rot fungus Schizophyllum commune

Mushroom-forming basidiomycetes colonize large areas in nature. Their hyphae are compartmentalized by perforated septa, which are usually covered by a septal pore cap (SPC). Here, we describe, for the first time, the composition and function of SPCs using the model system Schizophyllum commune. The SPC of S. commune was shown to consist of a proteinaceous matrix covered by a lipid membrane. The matrix was demonstrated to define the ultrastructure of the SPC and to consist of two main proteins, Spc14 and Spc33. Gene spc14 encodes a protein of 86 amino acids, which lacks known domain, signal or localization sequences. Gene spc33 encodes a 239 and a 340 amino acid variant. Both forms contain a predicted signal anchor that targets them to the ER. Immuno-localization showed the presence of Spc33 in the SPC but not in ER. From this and previous reports it is concluded that the SPC is derived from this organelle. Inactivation of spc33 resulted in loss of SPCs and the inability to close septa. The latter may well explain why vegetative growth and mushroom formation were severely reduced in strains in which spc33 was inactivated.

Septal pore complex morphology in the Agaricomycotina ( Basidiomycota) with emphasis on the Cantharellales and Hymenochaetales

The ultrastructure of septa and septum-associated septal pore caps are important taxonomic markers in the Agaricomycotina ( Basidiomycota, Fungi). The septal pore caps covering the typical basidiomycetous dolipore septum are divided into three main phenotypically recognized morphotypes: vesicular-tubular (including the vesicular, sacculate, tubular, ampulliform, and globular morphotypes), imperforate, and perforate. Until recently, the septal pore cap-type reflected the higher-order relationships within the Agaricomycotina. However, the new classification of Fungi resulted in many changes including revision of existing and addition of new orders. Therefore, the septal pore cap ultrastructure of more than 325 species as reported in literature was related to this new classification. In addition, the septal pore cap ultrastructures of Rickenella fibula and Cantharellus formosus were examined by transmission electron microscopy. Both fungi have dolipore septa associated with perforate septal pore caps. These results combined with data from the literature show that the septal pore cap-type within orders of the Agaricomycotina is generally monomorphic, except for the Cantharellales and Hymenochaetales. It appears from the fungal phylogeny combined with the septal pore cap ultrastructure that the vesicular-tubular and the imperforate type both may have arisen from endoplasmic reticulum. Thereafter, the imperforate type eventually gave rise to the perforate septal pore cap-type.

Septal Pore Cap Protein SPC18, Isolated from the Basidiomycetous Fungus Rhizoctonia solani , Also Resides in Pore Plugs

The hyphae of filamentous fungi are compartmentalized by septa that have a central pore. The fungal septa and septum-associated structures play an important role in maintaining cellular and intrahyphal homeostasis. The dolipore septa in the higher Basidiomycota (i.e., Agaricomycotina) are associated with septal pore caps. Although the ultrastructure of the septal pore caps has been studied extensively, neither the biochemical composition nor the function of these organelles is known. Here, we report the identification of the glycoprotein SPC18 that was found in the septal pore cap-enriched fraction of the basidiomycetous fungus Rhizoctonia solani. Based on its N-terminal sequence, the SPC18 gene was isolated. SPC18 encodes a protein of 158 amino acid residues, which contains a hydrophobic signal peptide for targeting to the endoplasmic reticulum and has an N-glycosylation motif. Immunolocalization showed that SPC18 is present in the septal pore caps. Surprisingly, we also observed SPC18 being localized in some plugs. The data reported here strongly support the hypothesis that septal pore caps are derived from endoplasmic reticulum and are involved in dolipore plugging and, thus, contribute to hyphal homeostasis in basidiomycetous fungi.

Enrichment of perforate septal pore caps from the basidiomycetous fungus Rhizoctonia solani by combined use of French press, isopycnic centrifugation, and Triton X-100

Septal pore caps occur in many filamentous basidiomycetes located at both sides of the dolipore septum and are at their base connected to the endoplasmic reticulum. The septal pore cap ultrastructure has been described extensively by the use of electron microscopy, but its composition and function are not yet known. To enable biochemical and functional analyses in the future, we here describe an enrichment method for perforate septal pore caps from Rhizoctonia solani. Our method is based on the combined use of French press and isopycnic centrifugation, using a discontinuous sucrose gradient followed by a treatment with Triton X-100. Enrichment was monitored by the use of scanning electron microscopy and transmission electron microscopy. Using the same isolation method, smaller septal pore caps were isolated from two other basidiomycetes as well. Furthermore, we showed pore-occluding material co-purified with the septal pore caps. This observation supports the hypothesis that septal pore caps play a key role in the plugging process of the septal pores in filamentous basidiomycetes.

Septal pore apparatus and nuclear division of Auriscalpium vulgare

Ultrastructure of the septal pore apparatus and nuclear division of Auriscalpium vulgare (Russulales) was examined with freeze substitution and is presented for inclusion in the AFTOL Structural and Biochemical Database (http://aftol.umn.edu). Previously unreported septal characters for the Russulales (Agaricomycotina) were observed: Septa of the hymenophore had bell-shaped perforated septal pore caps that may extend along the septum and a zone of organelle exclusion surrounded the septal pore apparatus. Metaphase I of meiosis and metaphase of mitosis were similar. Globular spindle pole bodies with electron-opaque inclusions were set within polar fenestrae of the nuclear envelope. The nuclear envelope was mostly intact with occasional gaps. Fragments of endoplasmic reticulum were present near the spindle pole bodies but did not form a polar cap. Structural characters may distinguish one or more clades of the Agaricomycotina and provide additional signal in phylogenetic analyses.

Septal pore apparatus and nuclear division of Auriscalpium vulgare

Ultrastructure of the septal pore apparatus and nuclear division of Auriscalpium vulgare (Russulales) was examined with freeze substitution and is presented for inclusion in the AFTOL Structural and Biochemical Database (http://aftol.umn.edu). Previously unreported septal characters for the Russulales (Agaricomycotina) were observed: Septa of the hymenophore had bell-shaped perforated septal pore caps that may extend along the septum and a zone of organelle exclusion surrounded the septal pore apparatus. Metaphase I of meiosis and metaphase of mitosis were similar. Globular spindle pole bodies with electron-opaque inclusions were set within polar fenestrae of the nuclear envelope. The nuclear envelope was mostly intact with occasional gaps. Fragments of endoplasmic reticulum were present near the spindle pole bodies but did not form a polar cap. Structural characters may distinguish one or more clades of the Agaricomycotina and provide additional signal in phylogenetic analyses.

Laser microdissection of fungal septa as visualised by scanning electron microscopy

Laser microdissection has been proven a successful technique to isolate single cells or groups of cells from animal and plant tissue. Here, we demonstrate that laser microdissection is suitable to isolate subcellular parts of fungal hyphae. Dolipore septa of Rhizoctonia solani containing septal pore caps were cut by laser microdissection from sections of mycelium and collected by laser pressure catapulting. Subsequently, microdissected septa were visualised using a wheat germ agglutinin labelling of cell walls, septa and septal pore caps and scanning electron microscopy. The use of laser microdissection on fungal cells opens new ways to study subcellular fungal structures and the biochemical composition of hyphal cells.