Showing papers on "Glutaraldehyde published in 1977"
TL;DR: The properties of a new fluorescence histochemical method for arylethylamines based on reaction with a mixture of 4% formaldehyde and 0.5% glutaraldehyde in aqueous solution are described, which produced a well-localized fluorescence reaction in tissues that was sufficiently intense for fine terminal noradrenergic axons to be seen.
Abstract: The properties of a new fluorescence histochemical method for arylethylamines based on reaction with a mixture of 4% formaldehyde and 0.5% glutaraldehyde in aqueous solution are described. At room temperature the aldehyde mixture produced a well-localized fluorescence reaction in tissues, which, when examined microscopically in aqueous solution, was sufficiently intense for fine terminal noradrenergic axons to be seen. If the tissue was subsequently dried, the fluorescence intensity increased. At the same time as inducing the fluorophores, the aldehyde mixture fixed the tissue to a standard well suited for electron microscopy. It thus proved possible to locate amine containing cells in the fluorescence microscope and subsequently examine their ultrastructure. In aqueous models, the aldehyde mixture formed fluorescent products with adrenaline, noradrenaline, dopamine, dopa, 5-hydroxytryptamine and 5-hydroxytryptophan, but not with histamine or octopamine. The fluorescence induced in the aldehyde mixture remained stable if the tissue was subsequently transferred to saline or distilled water and when it was dehydrated in ethanol and cleared with xylene, benzene, chloroform or acetone.
187 citations
01 Dec 1977
TL;DR: The study of glucose fermentation in immobilized cells shows that immobilization modifies the metabolism, while a slowing down of the cell's activity follows covalent binding.
Abstract: Porous glass beads are used to immobilizeSaccharomyces carlsbergensis cells. If silica pores are large enough, adsorption occurs. On the other hand, activation of the silica by glutaraldehyde allows the cells to bind onto the support. In the other case, the most porous glass has the greatest retention capacities. In all cases, 15 min is sufficient for support saturation by microorganisms.
131 citations
TL;DR: Quantitative comparison of photodichroism with mathematical models indicates that the pigment absorption vectors are aligned within +/- 50 degrees of the microvillar axes and are tilted into the surface of the membrane at an average value of about 20 degrees, suggesting that the bifunctional reagent quiets some molecular motion that is present after treatment with formaldehyde.
Abstract: Individual, isolated rhabdoms from dark-adapted crayfish (Orconectes, Procambarus) were studied with a laterally incident microbeam that could be placed in single stacks of microvilli. Concentration gradients of metarhodopsin along the lengths of microvilli were produced by local bleaches, accomplished by irradiation with small spots of orange light at pH 9 in the presence of glutaraldehyde or formaldehyde. No subsequent redistribution of pigment was observed in the dark, indicating an absence of translational diffusion. On the basis of comparison with other systems, glutaraldehyde, but not formaldehyde (0.75%), would be expected to prevent diffusion of protein in the membrane. Under the same conditions photodichroism is observed, indicating an absence of free Brownian rotation. Photodichroism is larger in glutaraldehyde than in formaldehyde, suggesting that the bifunctional reagent quiets some molecular motion that is present after treatment with formaldehyde. Quantitative comparison of photodichroism with mathematical models indicates that the pigment absorption vectors are aligned within +/- 50 degrees of the microvillar axes and are tilted into the surface of the membrane at an average value of about 20 degrees. The photoconversion of rhodopsin to metarhodopsin is accompanied by an increase in molar extinction of about 20% at the lambda maxand a reorientation of the absorption vector by several degrees. The transition moment either tilts further into the membrane or loses some of its axial orientation, or both. The change in orientation is 3.5 time larger in formaldehyde than in glutaraldehyde.
130 citations
TL;DR: Kappa -carrageenan was most suitable for immobilization of microbial cells and the glucose isomerase activity became stable for a long period even in the absence of gel-inducing reagents.
Abstract: To find a suitable matrix for immobilization of microbial cells, synthetic and natural polymers were screened. As a result,kappa-carrageenan,iota-carrageenan, furcellaran, sodium alginate, ethyl succinylated cellulose, succinylated zein, and 2-methyl-5-vinyl-pyridine-methylacrylate-methacrylic acid copolymer were studied. These polymers were induced to gel under mild conditions.Streptomyces phaeochromogenes cells having glucose isomerase activity were successfully immobilized in these polymer matrices. If a gelinducing reagent were added to a substrate solution, these gel matrices could be stabilized. The microbial cells did not leak out from the gel lattice. When these immobilized cells were treated with hardening reagents such as glutaraldehyde or tannins, the gel matrices were strengthened, and the glucose isomerase activity became stable for a long period even in the absence of gel-inducing reagents. Among these polymer matrices tested,kappa -carrageenan was most suitable for immobilization of microbial cells.
116 citations
TL;DR: Bismuth salts on aldehyde fixed tissue give a highly selective pattern of staining suitable for light and electron microscopy, and it is presumed that most bismuth staining of tissues is due to reaction with amine groups and is glutaraldehyde sensitive and some may be due to guanidine groups which are less sensitive to fixation by glutARaldehyde.
Abstract: Bismuth salts on aldehyde fixed tissue give a highly selective pattern of staining suitable for light and electron microscopy. Structures stained include the nucleolus, ribosomes, inter- and perichromatin granules, the Golgi complex beads and the outer face of the tubule doublets of mouse sperm, certain neurosecretory vesicles believed to contain biogenic amines, some junctions (some central synapses, neuromuscular junctions, tight junctions), specialized membranes such as the post acrosomal dense lamina of mouse sperm and the inner alveolar membrane of Paramecium , and a variety of structures associated with the cytoplasmic face of membranes, such as plasma membrane plaques, cleavage furrows, the leading edge of the spreading acrosome and sperm annuli. Staining is not reduced by nucleases and spot tests show no reaction between nucleic acids and bismuth under conditions similar to those used to stain tissues. However, spot tests do show strong binding of bismuth by basic proteins and by some phosphorylated molecules. It is hypothesized that bismuth reacts with cell components in two ways, distinguishable by their glutaraldehyde sensitivity. For example, staining of the nucleolus and ribosomes is blocked by glutaraldehyde but the inter- and perichromatin granules and the GC beads are unaffected. Spot tests show that basic proteins (histones, protamines, polylysine and polyargenine) and other molecules with free amino groups (5HT, tryptamine, dopamine) bind bismuth strongly, a reaction that is blocked to varying degrees by glutaraldehyde. We presume that most bismuth staining of tissues is due to reaction with amine groups and is glutaraldehyde sensitive and some may be due to guanidine groups which are less sensitive to fixation by glutaraldehyde. Organic phosphates may be the cause of the glutaraldehyde insensitive staining since ATP and some other phosphates bind bismuth in a reaction that is not blocked by glutaraldehyde.
79 citations
TL;DR: Improvements in the chemistry and chemical stability of stabilized alkaline 2% glutaraldehyde solution allow SGS to be continuously used for 14 days in situations of high dilution, or 28 days in situation of low dilution.
Abstract: The properties of stabilized alkaline 2% glutaraldehyde solution (SGS) are discussed. SGS is discussed with regard to its chemistry, antimicrobial properties, organic soil resistance, toxicity, corrosivity and chemical stability. SGS retains the maximum antimicrobial activity of alkaline glutaraldehyde solutions and the chemical stability heretofore observed only with acidic glutaraldehyde solutions. These improvements, along with the inherent resistance of glutaraldehyde to neutralization by organic soil, allow SGS to be continuously used for 14 days in situations of high dilution, or 28 days in situations of low dilution.
66 citations
TL;DR: In principle, enzyme-loaded erythrocytes can be used as a vehicle for enzyme replacement therapy in lysosomal storage diseases and glutaraldehyde treatment renders them more resistant to lysis without inactivating the enzymes that have been entrapped inside them.
47 citations
TL;DR: Fixation by immersion of skeletal muscle with a combination of 8% tannic acid and 2.5% glutaraldehyde revealed a prominent staining of the junctional “T” system, cell surface and lamellar structures at the cell periphery in undamaged muscle cells, which may indicate the presence of a special pool of proteins at this site.
37 citations
TL;DR: The milk‐clotting enzyme pepsin was immobilized onto beads of alumina, titania, glass, stainless steel, iron oxide, and Teflon for treating skim milk in a fluidized‐bed reactor and two covalent attachment procedures and two adsorption procedures were evaluated.
Abstract: The milk-clotting enzyme pepsin was immobilized onto beads of alumina, titania, glass, stainless steel, iron oxide, and Teflon for treating skim milk in a fluidized-bed reactor. Two covalent attachment procedures using silanized supports and glutaraldehyde and two adsorption procedures were evaluated. The three best catalysts were titania and glass, using the covalent attachment procedure, and alumina, using the adsorption procedure at pH 1.2. The pepsin adsorbed on alumina catalyst has commercial potential compared to the previously used glass catalyst. Attempts to increase the stability of pepsin adsorbed on alumina by cross-linking with glutaraldehyde were unsuccessful owing to the low pH necessary for optimum pepsin adsorption; Desorption of pepsin from alumina during reactor operation was determined. Regeneration of spent catalysts was only partially successful.
29 citations
TL;DR: Ruthenium red in glutaraldehyde and in osmium tetroxide, used for staining animal materials, is useful also with plant materials as discussed by the authors, and it was possible to observe that several cell sites are selected by R-Red staining.
29 citations
TL;DR: A double fixation method of preparing platelet suspensions for both scanning and transmission electron microscopy is outlined and prefixation in 0.1% glutaraldehyde allows for immediate preservation of morphologic characteristics induced by experimental procedures, but does not completely destroy platelet surface stickiness.
Abstract: A double fixation method of preparing platelet suspensions for both scanning and transmission electron microscopy is outlined. Prefixation in 0.1% glutaraldehyde allows for immediate preservation of morphologic characteristics induced by experimental procedures, but does not completely destroy platelet surface stickiness. Preservation of surface stickiness allows subsequent production of a platelet pellet for processing for transmission electron microscopy. This pelleting cannot be achieved when higher initial concentrations of glutaraldehyde are used for prefixation. Prefixation in 0.1% glutaraldehyde is also an appropriate initial step for preservation of platelets in suspension for scanning electron microscopy.
Patent•
15 Feb 1977
TL;DR: In this paper, superoxy-anion-forming metals, such as chromium, manganese, and the like, are removed from materials containing the same by contacting the material with an anionically-modified, nitrogen-containing polymer.
Abstract: Superoxy-anion-forming metals, such as chromium, manganese, and the like, are removed from materials containing the same by contacting the material with an anionically-modified, nitrogen-containing polymer, such as sulfite-modified, sulfate-modified, or chloride-modified chitosan.
TL;DR: The particle distribution of the intact RBC membrane depended on the use of glutaraldehyde fixation and glycerol cryoprotection and the lectin phytohemagglutinin induce particle rearrangement in intact cells.
Abstract: Freeze-fracture combined with quantitative electron microscopy of the intact human erythrocyte (RBC) and ghost revealed significant differences in their intramembranous particle coefficients External (E) fracture-faces of unfixed ghost membranes were found to contain 40% fewer particles than those of intact unfixed RBC The particle distribution of the intact RBC membrane depended on the use of glutaraldehyde fixation and glycerol cryoprotection Whereas glutaraldehyde- and glycerol-treated cells disclosed 70% fewer E-face particles than did intact unfixed cells, poly-L-lysine-treated, intact, unfixed RBC showed no such differences Treatment with a combination of poly-L-lysine and glutaraldehyde, however, increased the amount of E-face particles while reducing those of the protoplasmic (P) face The poly-L-lysine effect varied with its concentration and was unaffected by previous application of neuraminidase Nor did the lectin phytohemagglutinin induce particle rearrangement in intact cells Our data demonstrate that the processes of glutaraldehyde fixation and glycerol cryoprotection modify the RBC membrane by decreasing the number of E-face particles present In addition, the combination of poly-L-lysine and glutaraldehyde alters the affinity of some particles for one half of the membrane, suggesting that in freeze-fractured RBC, chemical bonds formed at the extracellular surface of the membrane can influence particle partitioning
TL;DR: Aggregation tends to enhance the previously demonstrated effects of glutaraldehyde in that reactivity with human IgE antibody, and ability to induce IgE antibodies in the Bordetella pertussis-treated rat, are further reduced.
Abstract: Timothy pollen extracts have been reacted with glutaraldehyde under conditions leading to different degrees of aggregation of the product. Aggregation tends to enhance the previously demonstrated effects of glutaraldehyde in that reactivity with human IgE antibody, and ability to induce IgE antibody in the Bordetella pertussis-treated rat, are further reduced. Ability to induce IgG antibody with specificity for unmodified extract is substantially retained in all aggregated products.
TL;DR: The sensitivity of isolated plastocyanin to glutaraldehyde further supports the conclusion that glutARaldehyde inhibits at the plastOCyanin level and thereby induces a break between P700 and cytochrome f.
Abstract: Treatment of spinach chloroplasts with glutaraldehyde causes an inhibition in the electron transport chain between the two photosystems Measurements of O2 flash yields, pH exchange, and fluorescence induction show that the O2 evolving apparatus, photosystem II and its electron acceptor pool are not affected The behavior of P700 indicates that its reduction but not its oxidation, is severely inhibited Cytochrome f is still reducible by photosystem II but also slowly oxidizable by photosystem I The sensitivity of isolated plastocyanin to glutaraldehyde further supports the conclusion that glutaraldehyde inhibits at the plastocyanin level and thereby induces a break between P700 and cytochrome f
TL;DR: Proteins were stained (brilliant red) by using 0·5% Ponceau 2R (C.I. 16150) in sulphuric acid (pH 2) at 40°C on sections 1–2 μm thick of tissues fixed in glutaraldehyde‐osmium tetroxide and embedded in epoxy resins.
Abstract: SUMMARY
Proteins were stained (brilliant red) by using 0·5% Ponceau 2R (C.I. 16150) in sulphuric acid (pH 2) at 40°C on sections 1–2 μm thick of tissues fixed in glutaraldehyde-osmium tetroxide and embedded in epoxy resins. The staining is free from precipitates and does not require the removal of the embedding media.
Section bleaching with 1–2% periodic acid at 40°C for 30–90 min was also carried out in order to obtain a more brilliant colour of the proteins which may be photographed using black and white films.
TL;DR: Physical removal of the cell wall of yeast and other fungal cells, by rapid mixing of the cells with glass beads after preliminary fixation in glutaraldehyde or formalin, removes thecell wall barrier to fixation and/or infiltration of the Cells for electron microscopy.
Abstract: Physical removal of the cell wall of yeast and other fungal cells, by rapid mixing of the cells with glass beads after preliminary fixation in glutaraldehyde or formalin, removes the cell wall barrier to fixation and/or infiltration of the cells for electron microscopy. The technique has been used on a variety of fungal cells which have been “difficult” to fix for electron microscopy, and appears to have wide applicability.
TL;DR: The biochemical azoindoxyl assay represents a useful method for combined qualitative and quantitative studies of acid beta-galactosidase in comparison with the indigogenic, the metal precipitation and the simultaneous azocoupling reactions for the in situ detection of acidbeta-galactsidase.
Abstract: The determination of various reaction constants yields the following assay for the photometric evaluation of acid beta-galactosidase (measurement of the azoindoxyl dye at 540 nm after extraction with dimethylformamide or -acetamide): 1.5 mM 5-Br-4-Cl-3-indolyl-beta-D-galactoside (1 mg dissolved in 0.05 ml dimethylformamide) and 0.01-0.015 ml hexazotized p-rosaniline/ml in 0.1 M citric acid-phosphate buffer, pH 4. By means of this procedure it becomes evident that the activity of the enzyme differs considerably in various rat organs; NaCl does not influence acid beta-galactosidase. -- Similar results were obtained with the indigogenic method; indigo can be dissolved and measured photometrically as the azoindoxyl dye. The enzyme is suppressed by high concentrations of hexazotized p-roaniline to 50%; low concentrations do not inhibit; the same is true for ferricyanide-ferrocyanide employed in the indigogenic media. -- The effect of glutar- and formaldehyde on acid beta-galactosidase cannot be investigated with the azoindoxyl reaction since the azoindoxyl dye partially withstands extraction from fixed blocks of tissue. On the basis of the biochemical findings the azoindoxyl technique can be recommended for the histochemical demonstration of acid beta-galactosidase: 7.5 mg (1.5 mM) 5-Br-4-Cl-3-indolyl-beta-D-galactoside (dissolved in 0.25 ml dimethylformamide) and 0.05-0.15 ml hexazonium-p-rosaniline in 10 ml 0.1 M citric acid-phosphate buffer, pH 4. After incubation the sections can be treated with osmium tetroxide followed by dehydration and mounting in resins or can be mounted without prior osmification of the azoindoxyl dye in glycerin jelly. The osmium chelate resists treatment with organic solvents; the stability of the chelate depends on the concentration of hexazotized p-rosaniline. After fixation in glutaraldehyde or in a mixture of form- and glutaraldehyde acid beta-galactosidase can be exactly localized in the lysosomes of many rat organs. In comparison with the indigogenic, the metal precipitation and the simultaneous azocoupling reactions for the in situ detection of acid beta-galactosidase the azoindoxyl procedure is superior if fixed material is used; it is equivalent or inferior in connection with membrane technique. The biochemical azoindoxyl assay represents a useful method for combined qualitative and quantitative studies of acid beta-galactosidase.
TL;DR: A scanning electron microscopic study of the elastin residue of normal human aortas has been made and purity has been judged by amino acid analysis.
Abstract: A scanning electron microscopic study of the elastin residue of normal human aortas has been made. Purification has been achieved by successive salt or guanidine extraction and by autoclaving, sodium hydorxide or collagenase treatment to remove contaminating proteins. Purity has been judged by amino acid analysis. The cleanest specimens have been obtained with NaOH. The elastin residue has been fixed with glutaraldehyde and osmium tetroxide. The specimens were rendered conductive by osmium-thiocarbohydrazide treatment and examined with a Cambridge scanning electronmicroscope without coating.
TL;DR: An efficient method of protein-protein conjugation yielding primarily monoconjugates is described, applicable if one of the two involved proteins is a glycoprotein.
TL;DR: The experimental results do not support the view that glutaraldehyde fixation, in the long term,attenuates the antigenic properties of proteins.
TL;DR: Fungicidal determinations resulted in a 99·99% reduction in viable count after 90 min contact with 0·5% (w/v) alkaline glutaraldehyde, and a considerable drop in spore production was also observed after treatment with alkaline glutamate.
Abstract: Fungistatic data were obtained from measurements of mycelial growth of several fungal species in the presence of acid or alkaline glutaraldehyde. Alkaline glutaraldehyde in concentrations > 0·1% (w/v) prevented growth of all species examined while 0·5% (w/v) acid glutaraldehyde was necessary to achieve this effect. Further fungistatic data were obtained using a Coulter Counter method to measure spore swelling. Fungicidal determinations resulted in a 99·99% reduction in viable count after 90 min contact with 0·5% (w/v) alkaline glutaraldehyde. A considerable drop in spore production was also observed after treatment with alkaline glutaraldehyde.
TL;DR: The surface disinfectant effect of glutaraldehyde in the gas-aerosol phase was investigated at different relative humidities and temperatures as discussed by the authors, and it was found that at a gasaerosols concentration of 15 to 20 mg/m3 and a relative humidity of about 80%, glut-araldehyde had a good disinfectant ability against both vegetative bacteria (decimal reduction time, less than 5 min) and bacterial spores (decal reduction time less than 45 min).
Abstract: The surface disinfectant effect of glutaraldehyde in the gas-aerosol phase was investigated at different relative humidities and temperatures. At a gas-aerosol concentration of 15 to 20 mg/m3 and a relative humidity of about 80%, glutaraldehyde had a good disinfectant effect against both vegetative bacteria (decimal reduction time, less than 5 min) and bacterial spores (decimal reduction time, less than 45 min). In spite of its low volatility, glutaraldehyde was more effective than formaldehyde when the two substances were compared on an "added amount" basis.
Journal Article•
TL;DR: Preliminary data indicate that with this protocol there is decreased background fluorescence and increased signal to noise ration in flow and nuclear and cytoplasmic fluorescence do not appear to vary significantly with respect to cell concentration or distribution on a slide or filter when cells are stained according to the glutaraldehyde protocol.
Abstract: A technique for Acridine Orange staining of cells for automated flow analysis involving post-staining fixation with Millonig's glutaraldehyde buffer is presented. Preliminary data indicate that with this protocol there is decreased background fluorescence and increased signal to noise ration in flow. In addition, stained cells may be stored for several months without alteration in cellular morphology or fluorescence characteristics. Finally, nuclear and cytoplasmic fluorescence do not appear to vary significantly with respect to cell concentration or distribution on a slide or filter when cells are stained according to the glutaraldehyde protocol.
Journal Article•
TL;DR: Cells treated with low concentrations of glutaraldehyde for a 10-sec interval were unable to incorporate 3H-leucine into TCA precipitable protein, respond to H-2 allogeneic cells in mixed lymphocyte reactions (MLR) and cell-mediated lympholysis (CML) assays, or display capping of cell surface immunoglobulin (Ig) with a fluoresceinated anti-Ig reagent.
Abstract: Cells treated with low concentrations of glutaraldehyde for a 10-sec interval were unable to incorporate 3 H-leucine into TCA precipitable protein, respond to H-2 allogeneic cells in mixed lymphocyte reactions (MLR) and cell-mediated lympholysis (CML) assays, or display capping of cell surface immunoglobulin (Ig) with a fluoresceinated anti-Ig reagent Such cells could stimulate and specifically block H-2 allogeneic CML activity but could not stimulate an H-2 allogeneic MLR response Trinitrobenzenesulfonic acid (TNBS) treated spleen cells were used to sensitize syngeneic splenocytes into displaying a cytotoxic effect against trinitrophenyl (TNP)-modified target cells Treatment of the stimulator cells with glutaraldehyde immediately after modification with TNBS did not impair their immunogenic activity Similar treatment of TNP-modified concanavalin A-stimulated lymphoblasts that were used as inhibitors in a CML cold target competition assay allowed such cells to retain their antigenicity Cells treated with glutaraldehyde before TNP-modification, however, were not antigenic in the cold target competition assay These data are compatible with TNBS acting on plasma membrane molecules directly to cause cells to be antigenic and immunogenic in the CML assay rather than affecting internal cellular components
TL;DR: No difference in Km values was observed between samples of catalase conjugates and untreated enzyme, either soluble or immobilized, and only 20% of the original glucose oxidase activity was lost while the untreated enzyme lost 90% of its activity under the same conditions.
Abstract: Soluble high molecular weight conjugates of catalase were prepared by intermolecular cross-linking with glutaraldehyde at pH 7.0. The same reagent was used to prepare high molecular weight soluble conjugates of glucose oxidase with and without catalase activity. For cross-linking of glucose oxidase the reactions were carried out at pH 4.5 in the presence of bovine serum albumin. No difference in Km values was observed between samples of catalase conjugates and untreated enzyme, either soluble or immobilized. When soluble glucose oxidase-catalase conjugates were incubated at 55°C for 18 min, only 20% of the original glucose oxidase activity was lost while the untreated enzyme lost 90% of its activity under the same conditions.
TL;DR: The production of an artifactual network in the nuclear sap of the salivary glands of Drosophila has been investigated and the inclusion of sucrose, and adenine nucleotides or other 'chelating' agents in the fixative reduces the effect, especially at low temperature.
Abstract: The production of an artifactual network in the nuclear sap of the salivary glands of Drosophila has been investigated. Mechanical stress to the cells, 3% glutaraldehyde containing more than 4 mM calcium, tannins, or cacodylate buffer or whose temperature is above 10 degrees C all enhance this artifactual effect. Over the range pH 6.65-7.4 there is no significant effect of pH. The inclusion of sucrose, and adenine nucleotides or other 'chelating' agents in the fixative reduces the effect, especially at low temperature (6-10 degrees C). These influences are additive and can abolish the artifactual effect. So too can 8% glutaraldehyde, but this is irrespective of temperature.
01 Jan 1977
TL;DR: In spite of its low volatility, glutaraldehyde was more effective than formaldehyde when the two substances were compared on an "added amount" basis.
Abstract: Received forpublication 1February 1977 Thesurface disinfectant effect ofglutaraldehyde inthegas-aerosol phase was investigated atdifferent relative humidities andtemperatures. Ata gas-aerosol concentration of15to20mg/m3anda relative humidity ofabout 80%,glutaraldehyde hada gooddisinfectant effect against bothvegetative bacteria (decimal reduction time,<5min)andbacterial spores (decimal reduction time,<45 min).Inspite ofitslowvolatility, glutaraldehyde was more effective than formaldehyde whenthetwosubstances were compared on an "added amount" basis.
TL;DR: It is proposed that the variation in pore diameters may indicate opening and closure of this orifice, and that the widely reported ‘central granule’ of the nuclear pore complex corresponds with the orifice in a closed configuration.
Abstract: SUMMARY
Aliquots of lymphocyte cell suspensions were pretreated according to the following three schedules before freeze fracturing: (a) prefixed with 2% glutaraldehyde before infiltration with 25% glyercol in medium RPMI-1640; (b) frozen in medium RPMI-1640 without additional pretreatment; and (c) frozen after pretreatment with 25% glycerol in medium RPMI-1640. The diameters of the fractured nuclear pore complexes of cells prefixed with glutaraldehyde were normally distributed within the range 70–120 nm (median 90 nm). The nuclear envelopes of 66–75% of cells processed through schedules b and c, which omitted glutaraldehyde fixation, had 70–120 nm diameter pores, while the remainder had pores with diameters in the range 120–175 nm. The large pores were structurally similar to the smaller pores except for their dimensions. These results indicate that glutaraldehyde gives rise to shrinkage of the larger pores to the minimum, smaller, diameter. Apparent orifices of at least 30 nm diameter were sometimes observed at the centres of these large pore complexes. We propose that the variation in pore diameters may indicate opening and closure of this orifice, and that the widely reported ‘central granule’ of the nuclear pore complex corresponds with the orifice in a closed configuration.