bluuebunny/crossref_metadata_2025 huggingface dataset:
Lactobacillus rhamnosus GG secreting an antigen and Interleukin‐2 translocates across the gastrointestinal tract and induces an antigen specific immune response
Authors: Kandasamy Matheswaran , Selvakumari Jayasurya Anita , Moochhala Shabbir , Huat Bay Boon , Kun Lee Yuan , Mahendran Ratha
Published: August 2011

Lactobacillus rhamnosus strain GG (LGG) is a probiotic organism. In this present study, LGG that express the green fluorescence protein (LGG‐GFP) and IL‐2 and GFP as a fusion protein (LGG‐IL‐2‐GFP) were used to examine bacterial uptake and the immune response induced by oral immunization. Using TEM to examine the intestinal tissue, the Lactobacilli were localized in M cells and in venules. After oral immunization, most of the bacteria were excreted in feces only a small fraction (0.15%) was retained in the intestine at 48 hr. However, more LGG‐IL‐2‐GFP was found in the MLN and spleen than LGG‐GFP. The loop ligation method was used to evaluate LGG uptake and both LGG‐GFP and LGG‐IL‐2‐GFP were found to translocate at the same rate. Analysis of LGG internalization in J774 macrophage cells indicated that IL‐2 increased survival of LGG and this may explain the increased presence of these bacteria in the MLN for a longer period. After oral immunization, specific mucosal antibody production as well as GFP specific CTL activity was demonstrated. IL‐2 co‐expression with GFP further enhanced antibody production and CTL activity. In conclusion, Lactobacillus rhamnosus GG expressing an antigen could generate an effective immune response to the antigen and IL‐2 improved the response generated probably by increasing LGG expressing antigen survival in immune cells.
A randomized experiment to examine unintended consequences of dietary supplement use among daily smokers: taking supplements reduces self‐regulation of smoking
Authors: Chiou Wen‐Bin , Wan Chin‐Sheng , Wu Wen‐Hsiung , Lee King‐Teh
Published: August 2011

Aims  We examined whether smokers' use of dietary supplements (e.g. vitamin C, multi‐vitamins) induces illusory invulnerability that in turn disinhibits smoking. Such supplement use may be perceived as conferring health credentials.

Design  A single‐factor (credentials: with or without) between‐subjects design was employed. Smokers were assigned randomly to take either a known placebo pill or a dietary supplement (in fact, the same placebo) in an ostensible health‐food test.

Setting  Laboratory at Kaohsiung Medical University, Taiwan.

Participants  Study 1 involved of a student sample consisting of 74 daily smokers, whereas study 2 involved a community sample consisting of 80 daily smokers.

Measurements  In Study 1, participants reported their perceived invulnerability following the manipulation. In study 2, pre‐ and post‐test measures of invulnerability were administered, and attitudes towards dietary supplements were assessed prior to the manipulation. In both studies, the dependent measure was the number of cigarettes smoked during completion of an unrelated survey.

Findings  Participants who believed that they were taking a dietary supplement smoked more cigarettes than did controls. Study 1 found support for the role of perceived invulnerability as a mechanism underlying this effect. Study 2 demonstrated the moderating effect played by attitudes towards dietary supplements: a more positive attitude towards supplements increased susceptibility to licensing effects.

Conclusions  Dietary supplement use may create illusory invulnerability, reducing the self‐regulation of smoking. Reminding health‐conscious smokers that multi‐vitamins do not prevent cancer may help such smokers to control their smoking and encourage them to stop.
A region on BTA14 that includes the positional candidate genes LYPLA1, XKR4 and TMEM68 is associated with feed intake and growth phenotypes in cattle1
Authors: Lindholm‐Perry A. K. , Kuehn L. A. , Smith T. P. L. , Ferrell C. L. , Jenkins T. G. , Freetly H. C. , Snelling W. M.
Published: August 2011

Feed cost for beef cattle is the largest expense incurred by cattle producers. The development of genetic markers to enhance selection of more efficient animals that require less feed while still achieving acceptable levels of production has the potential to substantially reduce production costs. A genome‐wide marker association approach based on the Illumina BovineSNP50 BeadChip™ was used to identify genomic regions affecting average daily feed intake (ADFI), average daily gain (ADG) and residual feed intake traits in a population of 1159 crossbred steers. This approach identified a region on BTA14 from 22.02 to 23.92 Mb containing several single‐nucleotide polymorphisms (SNPs) that have significant association with at least one of the traits. Two genes in this region, lysophospholipase 1 (LYPLA1) and transmembrane protein 68 (TMEM68), appeared to be logical positional and functional candidate genes. LYPLA1 deacylates ghrelin, a hormone involved in the regulation of appetite in the rat stomach, while TMEM68 is expressed in bovine rumen, abomasum, intestine and adipose tissue in cattle, and likely affects lipid biosynthetic processes. SNPs lying in or near these two genes were identified by sequencing a subset of animals with extreme phenotypes. A total of 55 SNPs were genotyped and tested for association with the same population of steers. After correction for multiple testing, five markers within 22.79–22.84 Mb, located downstream of TMEM68, and between TMEM68 and the neighbouring gene XKR4, were significant for both ADFI and ADG. Genetic markers predictive of feed intake and weight gain phenotypes in this population of cattle may be useful for the identification and selection of animals that consume less feed, although further evaluation of these markers for effects on other production traits and validation in additional populations will be required.
NUP98‐HOXA9‐transgenic zebrafish develop a myeloproliferative neoplasm and provide new insight into mechanisms of myeloid leukaemogenesis
Authors: Forrester A. Michael , Grabher Clemens , McBride Eileen R. , Boyd Ellen R. , Vigerstad Märta H. , Edgar Alexander , Kai Fui‐Boon , Da’as Sahar I. , Payne Elspeth , Look A. Thomas , Berman Jason N.
Published: August 2011

NUP98‐HOXA9 [t(7;11) (p15;p15)] is associated with inferior prognosis in de novo and treatment‐related acute myeloid leukaemia (AML) and contributes to blast crisis in chronic myeloid leukaemia (CML). We have engineered an inducible transgenic zebrafish harbouring human NUP98‐HOXA9 under the zebrafish spi1(pu.1) promoter. NUP98‐HOXA9 perturbed zebrafish embryonic haematopoiesis, with upregulated spi1expression at the expense of gata1a. Markers associated with more differentiated myeloid cells, lcp1, lyz, and mpx were also elevated, but to a lesser extent than spi1, suggesting differentiation of early myeloid progenitors may be impaired by NUP98‐HOXA9. Following irradiation, NUP98‐HOXA9‐expressing embryos showed increased numbers of cells in G2‐M transition compared to controls and absence of a normal apoptotic response, which may result from an upregulation of bcl2. These data suggest NUP98‐HOXA9‐induced oncogenesis may result from a combination of defects in haematopoiesis and an aberrant response to DNA damage. Importantly, 23% of adult NUP98‐HOXA9‐transgenic fish developed a myeloproliferative neoplasm (MPN) at 19–23 months of age. In summary, we have identified an embryonic haematopoietic phenotype in a transgenic zebrafish line that subsequently develops MPN. This tool provides a unique opportunity for high‐throughput in vivo chemical modifier screens to identify novel therapeutic agents in high risk AML.
Yield vs. Quality trade‐offs for wheat in response to carbon dioxide and ozone
Authors: Pleijel Håkan , Uddling Johan
Published: August 2011

Although it is established that there exist potential trade‐offs between grain yield and grain quality in wheat exposed to elevated carbon dioxide (CO2) and ozone (O3), their underlying causes remain poorly explored. To investigate the processes affecting grain quality under altered CO2 and O3, we analysed 57 experiments with CO2 or O3 exposure in different exposure systems. The study covered 24 cultivars studied in 112 experimental treatments from 11 countries. A significant growth dilution effect on grain protein was found: a change in grain yield of 10% by O3 was associated with a change in grain protein yield of 8.1% (R2 = 0.96), whereas a change in yield effect of 10% by CO2 was linked to a change in grain protein yield effect of 7.5% (R2 = 0.74). Superimposed on this effect, elevated CO2, but not O3, had a significant negative effect on grain protein yield also in the absence of effects on grain yield, indicating that there exists a process by which CO2 restricts grain protein accumulation, which is absent for O3. Grain mass, another quality trait, was more strongly affected by O3 than grain number, whereas the opposite was true for CO2. Harvest index was strongly and negatively influenced by O3, but was unaffected by CO2. We conclude that yield vs. protein trade‐offs for wheat in response to CO2 and O3 are constrained by close relationships between effects on grain biomass and less than proportional effects on grain protein. An important and novel finding was that elevated CO2 has a direct negative effect on grain protein accumulation independent of the yield effect, supporting recent evidence of CO2‐induced impairment of nitrate uptake/assimilation. Finally, our results demonstrated that processes underlying responses of grain yield vs. quality trade‐offs are very different in wheat exposed to elevated O3 compared with elevated CO2.
Increased quality and functionality of kimchi when fermented in Korean earthenware (onggi)
Authors: Jeong Ji‐Kang , Kim Young‐Wook , Choi Hye‐Sun , Lee Dong Sun , Kang Soon‐Ah , Park Kun‐Young
Published: August 2011

Kimchi was fermented in onggi and other containers at 4 °C for 4 weeks. Two types of onggi were used: glazed and nonglazed onggi. Other containers such as polyethylene plastic containers, polypropylene containers for use in a kimchi refrigerator, stainless steel and glass bottles were used. During the fermentation, kimchi fermented in onggi showed relatively stable acidity change compared with those in the other containers. Also it showed higher lactic acid bacteria multiplication (108–109 cfu g−1 at 4th week) but slower general aerobic bacteria multiplication than the others. The springiness of kimchi fermented in onggi was more than 50% at 4th week, it is higher than the others. Its carbonic acid taste and overall acceptability were excellent in sensory evaluations. The antioxidative and cancer‐cell antiproliferative activities were also greater in kimchi fermented in onggi than the others. Between the two types of onggi, nonglazed onggi showed better results in both properties and functionalities of kimchi. Onggi showed higher gas permeability than polyethylene plastic containers and glass bottles because of its porous structure, and lactic acid bacteria could have benefited from this condition. It possibly contributes to optimal ripening of kimchi resulting in better properties, especially texture and taste, and health functionalities.
Influence of gender and anaesthesia type on day surgery anxiety
Authors: Mitchell Mark
Published: August 2011

mitchell m. (2011) Influence of gender and anaesthesia type on day surgery anxiety. Journal of Advanced Nursing 68(5), 1014–1025.

Aim.  To investigate the possible influence of gender and anaesthesia type on anxiety prior to day surgery.

Background.  Elective surgery undertaken on a day, short stay or ‘day of surgery’ basis is growing and much emphasis also placed on ‘enhanced recovery’ for in‐patient surgery. During such brief episodes preoperative apprehension can be considerable but the opportunity to help reduce anxiety is minimal and formal plans uncommon.

Method.  As part of a larger study, a questionnaire was distributed to 1606 patients undergoing day surgery, with anaesthesia (2005–2007). Participants were requested to return the questionnaire by mail 24–48 hours following surgery, with 674 returned. Data were analysed using descriptive statistics and multivariate analysis of variance.

Results.  Of the total patients 82·4% experienced anxiety on the day of surgery with the wait, anaesthesia and possible pain being common anxiety‐provoking aspects. The majority preferred to receive information between 1–4 weeks in advance and participants experiencing general anaesthesia required information at a statistically significantly earlier stage. General anaesthesia patients were statistically significantly more anxious than local anaesthesia patients and desired more information. Female patients were statistically significantly more anxious, anxiety commenced earlier and they preferred to wait with a relative/friend or talk with other patients.

Conclusions.  Anxiety was experienced by the majority of participants but was more prevalent amongst general anaesthesia and female patients. For general anaesthesia patients, a comprehensive level of information may be required a number of weeks prior to surgery and gender differences associated with the preoperative wait may require greater consideration.
Low‐contrast development of electron micrographs
Authors: Farnell G. C. , Flint R. B.
Published: August 2011

From the fact that the response of photographic materials conforms to the ‘single‐hit’ expression for exposure to electrons of the energies used in electron microscopy, it follows that the density log exposure curves have a shape substantially independent of the degree of development in conventional solutions. The only way in which the density range corresponding to a given ratio of intensities in the image can be reduced is by reduction of the exposure. The visibility of low‐intensity detail in the image is adversely affected, however. A developer formula is presented with which the density contribution per electron decreases with increasing exposure so that contrast is less than if determined solely by the nature of the exposure. The effectiveness of the solution is demonstrated and practical recommendations are made regarding its use.
The structure and formation of the egg membranes in Nasonia vitripennis (Walker) (Hymenoptera, Pteromalidæ)
Authors: Richards J. G.
Published: August 2011

A histochemical and fine structural study of the vitelline membrane and chorion was carried out. Following the uptake of protein at the oöcyte plasma membrane during yolk synthesis, the egg membranes are laid down in sequence in the intercellular space between the oöcyte and the follicle cells. The follicular epithelial cells alone contribute towards their formation. The structure of the chorion is much simpler than that of Bombus, the only other hymenopteran previously investigated.
The effect of refractive index of the mounting medium on the apparent diameter of latex particles and glass fibres
Authors: Skene Donald S.
Published: August 2011

The diameters of latex particles were measured in different media with a light microscope. The microscope was fitted with a filar micrometer eyepiece and, for measurement, the cross‐wire was set on the centre of the dark line forming the edge of the image. Electron microscopy was used for comparison. Good agreement between light and electron microscopy was obtained using media of relative refractive index (nd object/nd medium) between 1·06 and 1·08. With higher relative refractive indices the light microscope underestimates dimensions for particles 4·17 μ diameter and overestimates for particles 2·7 μ diameter.
Electron phase and amplitude images of stained biological thin sections
Authors: Haydon Glen B.
Published: August 2011

Very‐high‐resolution electron microscopy demonstrates similar phase image ‘particles’ in uncontaminated, unsupported thin sections of epoxy‐embedded biological material and in simple thin carbon films, both of which show better than 5 Å resolution. Under such conditions it is possible to examine biological specimens for amplitude images well below the 20 Å limit commonly accepted for such preparations. With this very high resolution discrete amplitude images are found in sections after treatment with heavy‐metal stain. These amplitude images are as small as 10 Å, a size equal to that of lead aggregates, which they may represent.
Comparative studies on the ultrastructure of the thyroid parafollicular C‐cells
Authors: Welsch U. , Flitney E. , Pearse A. G. E.
Published: August 2011

A comparative study of the fine structure of the parafollicular C‐cells in the thyroid glands of two rodents (guinea‐pig and mouse), and a primitive primate (Tupaia glis), shows a large number of similarities with the previously reported findings of several authors.

In two of the three species the C‐cells possess numerous long cytoplasmic processes and they are preferentially elongated or ovoid in shape. This accords with their optical‐microscopic appearance in pig thyroid, but the feature appears not to have been recorded previously at electron‐microscope level.

The finding of ‘dark’ and ‘light’ cells is confirmed and it is suggested that the dark cell represents a different (degenerative) physiological state. The electron‐microscope appearance, however, may be due partly to differential retention of cytoplasmic components (including possibly biogenic amines), and hence they may be partly fixation artifact.

Other notable features of the C‐cells, such as microtubules and cilia, are recorded and discussed. The primary importance of fixation is particularly stressed.
The surface structure of mouse peritoneal cells—a study with the scanning electron microscope
Authors: Carr Ian , Clarke J. A. , Salsbury A. J.
Published: August 2011

Normal and stimulated peritoneal cells have been examined with the Stereo‐scan scanning electron microscope. Normal lymphocytes were identified by their small size; some had numerous small stubby or pointed processes on their surfaces. Macrophages were larger and had ridge‐like processes. Many of the stimulated cells showed no significant deviation from normal. Others, all macrophages, showed a marked deviation from the normal spherical shape. Ridge‐like processes were more prominent on their surfaces, and large flange‐like processes and a few finger‐like processes were seen. This suggests that on stimulation peritoneal cells become more deformable and therefore irregular in shape, and that their surfaces become rougher.
Histological control of specimen selection for electron microscopy
Authors: Tarin D. , Spencer A. T.
Published: August 2011

This technique enables small specimens for electron microscopy to be obtained from specific zones of large pieces of tissue. The method depends on the use of frozen sections, for localization of histological features, but circumvents the risk of ice‐crystal damage to the specimens selected.
An electron‐microscopic study of cholinesterase distribution in the rat adrenal medulla
Authors: Lewis P. R. , Shute C. C. D.
Published: August 2011

The distribution of cholinesterase activity in the rat adrenal medulla was studied after initial fixation with glutaraldehyde, which preserves the osmiophilia of the noradrenaline‐containing cells. Staining for true cholinesterase activity was associated with all the identifiable elements of the preganglionic sympathetic innervation. The morphology and enzyme staining of the endings on the chromaffin cells was similar to that seen in cholinergic synapses in the central nervous system. The endings contained occasional dense‐cored vesicles and the possible significance of such vesicles at an undoubted cholinergic synapse is discussed. Staining for pseudocholinesterase activity, by contrast, was associated particularly with the Schwann cells and to a lesser extent with the chromaffin cells, but not with the axons and their processes.
A phase‐contrast device with positive and negative image contrast
Authors: Pluta M.
Published: August 2011

A phase‐contrast device comprising objectives provided with two concentric phase rings, one positive and one negative, and a condenser with aperture diaphragm provided with two concentric transparent rings conjugate with the phase rings and covered by two crossed polarizing films, is described. The change of image contrast from positive to negative, and vice versa, is obtained by rotation of a polarizer placed before the condenser or above the objective. Properties and advantages of the device are discussed.
A simple microtome capable of cutting sections of plastic‐embedded material down to 1 μm in thickness
Authors: Magill H. , Gunning B.
Published: August 2011

Details of the design of an easily constructed microtome are presented. The instrument uses glass knives to cut sections down to about 1 μm in thickness, and is suitable for the examination of plastic‐embedded material.
A clamp to hold electron‐microscope grids for shadowing
Authors: Gregory David W.
Published: August 2011

The construction and use of a clamp to hold electron‐microscope grids for shadowing is described. The clamp holds up to twelve grids at one time and is particularly useful for undergraduate teaching.
The refractive index and dry mass distribution of mammalian erythrocytes
Authors: Thaer A.
Published: August 2011

Quantitative microscopical methods are now of outstanding importance in the diagnosis and assessment of mammalian red blood cells.

After release from the erythropoetic organs, all red blood cells in the peripheral blood of mammals pass through the reticulocyte stage before becoming mature erythrocytes. Therefore, the determination of reticulocyte number is used as an index of erythropoetic activity in clinical hæmatology.

The corpuscular solid concentration, the corpuscular dry weight, the osmotic resistance and the size of red blood cells can be determined by use of microscopical methods. For the determination of the corpuscular solid concentration of erythrocytes, cell refractometry by the use of phase‐contrast microscopy has proved to be a very sensitive and accurate method. The dry weight of single erythrocytes may likewise be obtained by the use of interference microscopy with relatively high accuracy. Both of these methods might also be of use in clinical hæmatology.

In the experimental work described in this paper, both methods mentioned above are used to answer the question: whether and under what conditions the refractive index as a criterion of measurement of the solid concentration and the relative dry weight can be determined separately for reticulocytes and erythrocytes from the same blood sample by use of microscopical methods alone. The frequency distributions of refractive indices and relative dry weights determined on human reticulocytes and mature erythrocytes show significantly lower mean values for the reticulocyte generation in accordant with findings published by other authors and obtained by use of sometimes entirely different methods. The additional information gained by the determination of the frequency distribution of refractive index and dry weight of reticulocytes and mature erythrocytes separately in a blood sample and the necessity for simplifying and automating such methods for hæmatological diagnosis are discussed.
Studies on insect fibrous proteins: the structural protein of the ootheca in the praying mantis, Sphodromantis centralis Rehn
Authors: Kenchington W. , Flower N. E.
Published: August 2011

Histological studies have been made on the left colleterial gland which secretes the α‐type structural protein in the ootheca of Sphodromantis centralis. Three distinct types of secretory cell are present in the gland wall, and each has a complex apical ‘end‐apparatus’ composed of closely packed canalicules radiating from a central cavity. Structural protein is synthesized in the cytoplasm, passed through the canalicules into the cavity of the end‐apparatus, and extruded into the lumen as strongly birefringent, apparently membrane‐free globules. At this stage the globules consist of numerous concentric layers of long, parallel fibrils, 0·05–0·1 μm in diameter. In each successive layer the fibril direction is rotated through an angle of about 18° about an axis perpendicular to the layers, in the manner of cholesteric phase systems. When sectioned, the globules reveal a regular ‘lamellated’ structure similar to that apparent in sections of some insect and crustacean cuticles. A three‐dimensional model has been constructed to illustrate this phenomenon. During ootheca formation fibrils become transformed into thin ‘crystalline’ ribbons, about 15 μm long, 1–2 μm wide and 200–300 Å thick. Very regular diamond‐shaped ribbons may be obtained in vitro by mixing structural protein globules from dissected left glands with optimum concentrations (0·005–·025 m) of calcium chloride in unbuffered solution. Experiments suggest that calcium ions play an important part in the natural process of ribbon formation.
Observations on the fine structure and development of the spindle at mitosis and meiosis in a marine centric diatom (Lithodesmium undulatum)
Authors: Manton I. , Kowallik K. , Stosch H. A. von
Published: August 2011

A spindle precursor is shown to be a relatively constant feature in non‐dividing cells of Lithodesmium undulatum and its structure is described and illustrated. It is a small somewhat rectangular body closely pressed to the nuclear envelope and composed of a series of parallel plates of semi‐opaque material placed edgeways on a square ground plan about 0·6 μm in length and breadth. The plates at each end are the densest and the next most dense marks the centre. During prophase in the spermatogonia the spindle proper is laid down between the precursor and the underlying nuclear envelope. It consists of tubules which increase in size and number as prophase advances, the spindle poles being marked by extensions from the terminal plates of the precursor, the rest of which ultimately breaks down, thereby permitting rapid elongation of the spindle itself to take place. After breakdown of the nuclear envelope, the spindle sinks into the mass of chromosomes, as previously described with the light microscope in other diatoms. The fine structure of the metaphase spindle is described and illustrated, special attention being paid to the number and arrangement of the component microtubules, both in a normal cell and a giant cell. Anaphase is not examined but a few stages of telophase are added, including some stages of cleavage. The results are discussed in a preliminary way pending completion of a study of meiosis. Points raised include consideration of a possible homologue of the so‐called centrosome of pennate diatoms and certain aspects of the numerical relation between chromosomes and spindle tubules.
A microspectrofluorimeter with epi‐illumination and photon counting
Authors: Pearse A. G. E. , Rost F. W. D.
Published: August 2011

This paper describes an instrument for microspectrofluorimetry, based on the Leitz Microspectrograph. Important features are monochromatic epi‐illumination and photon counting.
A sandwich‐embedding technique for monolayers of cells cultured on Araldite
Authors: Smith W. , Gray E. W. , Mackay J. M. K.
Published: August 2011

A method of culturing cell monolayers on Araldite to facilitate selection and sectioning of specific cells is described. The method was originally developed for the selection of virus‐induced multi‐nucleate cells, but is applicable to other systems.
Endocytosis: The different energy requirements for the uptake of particles by small and large vesicles into peritoneal macrophages
Authors: Casley‐Smith J. R.
Published: August 2011

The uptake of various, different‐sized particles by macrophages was studied using the electron microscope. In addition to observing normal cells, cells were examined which had been inhibited by exposing them to low temperature (4°C), and to a number of metabolic poisons.

It was found that large particles (> 01 μm) enter the cells and are contained in large vesicles (0·1–5 μm). Small particles (< 50 nm) may also enter the cells by this process. They enter most frequently, however, by passing into small (∼ 70 nm) vesicles. These may later coalesce and their contents adhere to give a second kind of large vesicle.

The various inhibitors prevented the ingestion of the large particles (and of the small particles en masse) into large vesicles, but did not prevent their initial adsorption onto the plasma membranes. They did not prevent nearly normal numbers of small particles from entering the pre‐existing small vesicles, nor their subsequent fusion into the second class of large vesicles.
Quantitative studies on the preservation of choline and ethanolamine phosphatides during tissue preparation for electron microscopy
Authors: Cope G. H. , Williams M. A.
Published: August 2011

The loss of 14C‐ethanolamine‐ and 3H‐choline‐labelled phospholipids from rat liver during preparation for electron microscopy by some less frequently used processing methods has been examined. Permanganate and formaldehyde‐potassium dichromate fixation followed by Araldite embedding were investigated and five procedures involving embedding in water‐miscible methacrylates (GMA). These procedures included a conventional method of dehydration and embedding in GMA, a low‐temperature GMA embedding method, dehydration with ethylene glycol, freeze drying and freeze substitution. These results are compared with those obtained after conventional tissue preparation (presented previously, Cope & Williams, 1969). Formaldehyde‐potassium dichromate compared favourably with the conventional procedures for the preservation of both phosphatides, especially phosphatidyl ethanolamine. Permanganate fixation was much less effective. Severe loss of both phosphatides occured after freeze drying and freeze substitution in glutaraldehyde‐alcohol. GMA is shown to be a more potent phospholipid solvent than ethanol under the conditions employed. Low‐temperature embedding reduced the loss of phosphatidyl choline during embedding. Results obtained by scintillation counting were confirmed by grain counts on thick‐section autoradiographs. No direct relationship between extraction and the electron‐microscopic appearance of membranes was discernible. It is believed that membrane prominence is largely dependent upon the electron density of the surrounding cytoplasm rather than on the degree of phospholipid extraction.
A stable iron‐hæmatoxylin solution for staining the chromatin of cell nuclei
Authors: Slidders W.
Published: August 2011

By using ferrous sulphate as mordant, sodium iodate as oxidant and aluminium chloride as stabilizer, it proved possible to formulate a stable single‐solution iron‐hæmatoxylin. This provides a resistant nuclear stain, sufficiently acid‐fast to be suitable for counterstaining with van Gieson or the trichrome methods. This solution, ferrous hæmatein, can replace the celestin blue‐hæmalum sequence.
A new acridine orange method for staining nucleic acid and its application to germinating wheat seeds
Authors: Price C. E.
Published: August 2011

A new method of nucleic acid staining based on acridine orange is described and applied to sections of wheat seed. The coleorhiza of the ungerminated seed has an RNA content as high as the embryonic leaves and roots. During the first 24 h of germination the RNA levels of the roots, leaves and scutellum remain relatively unchanged while that of the coleorhiza falls to a low level. The possibility is discussed that the rapidly‐mobilized RNA functions as a storage product which is hydrolysed and incorporated into fresh RNA in the adjacent embryo during germination.
The effect of glycerol on the structure of lecithin membranes; a study by freeze‐etching and X‐ray diffraction
Authors: Buckingham J. H. , Staehelin L. A.
Published: August 2011

The structure of bimolecular lecithin leaflets (or bilayers) has been studied in lecithin–water and lecithin–glycerol–water systems using freeze‐etch and X‐ray diffraction techniques.

The X‐ray diffraction experiments indicate that the thickness of these bilayers increases both with increasing lecithin concentration and with the addition of glycerol. Due to the glycerol‐induced thickening of the bilayers the limiting value of about 5 nm is reached at a lower lecithin concentration (70% lipid) in glycerol‐treated systems than in specimens prepared with pure water (75% lipid). It is suggested that the addition of glycerol increases molecular order within the bilayers by ‘cross‐linking’ the polar end‐groups of the lecithin molecules. The resultant straightening of the molecules increases the bilayer thickness for all concentrations below 75% lecithin.

The freeze‐etch studies show that the fracturing properties of the bilayers at −100°C change both when the concentration of lecithin increases from 1% to 80% and when glycerol is added to the preparations. The appearance of all the exposed membrane faces can be explained by a bilayer arrangement of the lecithin molecules. At concentrations below 75% lipid in lecithin‐water systems and concentrations below 70% lipid in glycerol‐treated specimens the membrane faces are usually covered with plaque structures, which represent island‐like remnants of the partly split away upper‐lying layer of the bilayers (Staehelin, 1968a). Above these concentrations most bilayers are completely split during the fracturing process. The freeze‐etch results also support the idea that glycerol strengthens the cohesive forces in the surface layers of the bilayer membranes and indicate that plaque formation depends on the presence of ‘free’ water between the bilayers.

A parallel freeze‐etch investigation of the hydration of bulk lecithin has shown that it could be a two‐step process. In the first stage the lecithin molecules appear to be arranged into bilayers while the bound water layer is formed. In the second stage the free water seems to penetrate as a sheet in between the bound water layers of the lipid bilayers and forces them apart.

Possible biological implications of these studies are briefly discussed.
The analysis of polarized light using a stereographic projection of the Poincaré sphere
Authors: Mallon H. D.
Published: August 2011

The mathematical analysis of a beam of polarized light into two constituent beams of plane‐polarized light vibrating at right angles to each other and with a difference of phase, can be somewhat time‐consuming. This paper describes a graphical method of solution, of reasonable accuracy, which can be carried out rapidly. The simple geometry of the representation of a polarized beam as a point on the surface of the Poincaré sphere is explained together with the stereo‐graphic projection of the surface of the sphere on a diametral plane. Examples are given of the use of the stereogram in conjunction with the polarizing microscope for the determination of phase difference of a transparent crystal, for the determination of the state of polarization after transmission through a bire‐fringent crystal and for the analysis of superimposed crystals.
A method for determining the fibrillar angle in wood tracheids
Authors: Page D. H.
Published: August 2011

The predominant direction of fibrils in isolated walls of wood tracheids can be determined from observation of the extinction position in the polarizing microscope. This method cannot be used directly to obtain the fibrillar direction for separate intact cells, because the light path must traverse the opposite walls of the cell, the fibrillar angles of which are similar but of opposite sense.

This difficulty may be overcome by impregnating the cell lumens with mercury and using incident (epi‐) illumination with crossed polars. The incident light now passes through the upper wall of each cell, suffers reflection at the mercury surface and returns through the same wall. The extinction position gives the predominant fibrillar direction of the upper wall.

The method is rapid, has been found suitable for tracheids in wood, wood pulp and paper, and should be of value for the study of other plant cells such as cotton.
Tritium‐labelled Araldite as an internal standard for quantitative autoradiography using the electron microscope
Authors: Flitney F. W.
Published: August 2011

A method is described for making quantitative measurements of radioactivity from electron‐microscope autoradiographs. Autoradiographs are made from sections of tissue which contain a seam of tritium‐labelled Araldite of known radioactivity. The latter serves as an internal standard and an estimate of the tissue activity is obtained by comparing the tissue grain density with that over the standard. The method can avoid many of the problems which are commonly encountered in making quantitative measurements of tissue radioactivity.
Image formation from alloys in the field‐ion microscope
Authors: Southworth H. N. , Ralph B.
Published: August 2011

Previous studies of solid‐solution alloys with the field‐ion microscope are reviewed, and it is pointed out how their image contrast must be determined by the different response of the atomic species towards field ionization and field evaporation. The former effect, termed selective ionization, describes the relative brightness of the image points arising from the different atomic species. It is pointed out that this process is secondary to considerations of selective evaporation since the latter determines whether both species can occupy potential imaging sites in any given region of the image.

Selective evaporation describes the relative fields required to evaporate the two species of atom from the specimen surface. In general the solute species will either be preferentially evaporated from the surface defined by the solvent species, or will be preferentially retained onto it. Consideration of the mechanism of selective evaporation shows that its degree may be expressed by a simple parameter calculated from known thermodynamic and atomic‐bonding data. In the later sections the model is applied to reported field‐ion data from solid‐solution alloys, excellent correlation being obtained.
Enhanced contrast in electron microscopy of unstained biological material
Authors: Johnson H. M. , Parsons D. F.
Published: August 2011

Electron‐optical methods of enhancing contrast are being investigated in order to eliminate the artifacts associated with heavy‐metal contrasting procedures. This report gives a quantitative discussion of the application of the strioscopic dark‐field method to model biological objects. Beam‐stopping apertures were constructed by microwelding fine platinum wires on to platinum objective apertures. These were precisely located and centred in the back focal plane of the objective lens. Contrast was evaluated by a sensitive micro‐Faraday cage system. Bright‐field and strioscopic contrast of organic layers up to 557 nm thickness were investigated and related to the transition from single to plural scattering. The spherical‐aberration phase‐contrast contribution could be separated from the amplitude contrast in bright‐field microscopy by extrapolation to zero object thickness. Strioscopic contrast of thin layers was found to be much greater than bright field aperture contrast and shown to be non‐linear with respect to thickness in the region 0·7–10 nm. For diffraction patterns made up of a few discrete beams the resolution is high, but is considerably less for diffusely scattering and thick objects. Strioscopy may be the method of choice for obtaining contrast of unstained or weakly‐stained objects at high acceleration voltage.
Preparation of animal tissues for surface‐scanning electron microscopy
Authors: Boyde A. , Wood C.
Published: August 2011

This paper surveys common problems associated with the preparation of animal tissues so that natural or artificial surfaces may be studied in the scanning electron microscope (SEM). Problems arise because (1) we need to prepare surfaces free from extraneous solids in solution (e.g. mucus, blood, tissue fluid); (2) surfaces of simple sections generally do not reveal significant information so that other methods of ‘opening up’ internal organization must be developed; (3) we need to remove the preponderant tissue component—water—without significantly altering the remaining structure, which generally has a very low density, is fragile and is non‐conducting.

In the case of natural, free surfaces to soft tissues, isolated cells or cultures the most useful approach seems to be (a) to wash with a suitable isotonic medium; (b) to fix with an aldehyde; (c) to harden with Parducz's (1967) OsO4, HgCl2 fixative.

Artificial surfaces revealing details of internal organization are best prepared by dissecting and ripping aldehyde‐perfused and fixed tissue; the duration of fixation has been found to influence the location of the ripping planes in CNS. Enzymatic digestion may usefully be combined with dissection, but must be conducted before fixation.

Wet tissue surface is least distorted following freeze drying from a non‐polar solvent (e.g. amyl acetate) which has substituted the tissue water. Critical‐point drying requires simple apparatus and is also generally satisfactory, but it does cause some bulk shrinkage.

Specimens are best coated with a combination of carbon and gold to render them conducting: carbon because it is easily scattered and thus tends to cover very rough surfaces, and because it is tough and stabilizes the surface; gold because it is the most convenient heavy metal having high secondary‐electron emission to evaporate.

Dried soft tissues have very low bulk densities, and beam‐penetration artifacts (bulk charging and edge artifacts) are therefore troublesome. These may be limited by working at as low an accelerating beam voltage as can be tolerated to achieve the required resolution. The preparation and examination of stereo‐pair micrographs is recommended as a routine because images can then be interpreted in terms of topography alone, and brightness variations of artifactual or unknown origin can be mentally excluded.
Staining of immature collagen in cattle
Authors: Schmitt O. , Dumoni B. L. , Dreyer J. H.
Published: August 2011

The staining of newborn calf tissues, using resorcin–fuchsin under different conditions, indicates that this dye is not suitable for studies of immature collagen in this species, but that a more promising stain is Van Giesons's picro–fuchsin.
An automatic sampling stage microscope for stereology
Authors: Weibel Ewald R.
Published: August 2011

A new instrument is described which permits automatic sampling of microscopic fields on sections. For this purpose a motor‐driven high‐precision specimen stage was fitted to a microscope. The motors are controlled by an electronic unit to permit the following modes of operation: (a) continuous operation by a four‐position joy‐stick switch for easy and rapid scanning of the preparation; (b) single‐step operation, where the specimen is displaced in either the x‐ or the y‐direction by a preselected distance adjustable between 15 μm and 20 mm; (c) automatically repeated step operation, where the steps occur at intervals pre‐selectable between 0·5 and 34 sec; (d) scanning operation, where the specimen continuously moves along the x‐axis and is shifted to the next line at a preselected distance when the end of the sweep is reached. The details of principle and operation are presented together with a few typical examples of application.
Ultracryotomy: a technique for cutting ultrathin sections of unfixed frozen biological tissues for electron microscopy
Authors: Hodson Stuart , Marshall John
Published: August 2011

An apparatus is described which facilitates the sectioning of unfixed and unembedded biological tissues for electron microscopy. The specimen is maintained at the temperature of boiling liquid nitrogen; the knife temperature may be controlled between −50 and − 150°C. Under the worst operating conditions specimen drift with respect to the knife is about 5 nm/sec, and suggestions are made on how this drift may be further reduced. We propose to call the apparatus an ‘ultracryotome’.

Electron micrographs of a variety of unfixed, unembedded and unstained tissues are shown and commented on. Some developments of experimental electron microscopy which may be expected to develop from use of the ultracryotome are listed.
Use of the X‐ray projection microscope in the examination of bone and cartilage sections
Authors: Rodriguez‐Garcia J. J. , Little K.
Published: August 2011

The conditions of use of the X‐ray projection microscope for the investigation of bone and cartilage sections have been examined. For the non‐calcified tissues an aluminium target operating at 6 kV has been found the most satisfactory. Aluminium at 15 kV and a copper target at 6 kV are also useful. The instrument is not so satisfactory as conventional microradiography for examination of fully calcified bone, but provides useful information about the softer tissues.

In the X‐ray projection microscope a beam of X‐rays produced as nearly as possible from a point is used as a source of illumination, and objects to be photographed are placed between, but not in contact with, the source of X‐rays and the photographic plate. All parts of the specimen or section are equally in focus, so that details from one surface to the other are recorded on the plate with equal sharpness. Because of this depth of focus stereoscopic photographs can be taken to give additional information.

The X‐ray microscope is capable of giving useful information where density differences between tissue components are of importance. The resolving power is governed by the size of the X‐ray source, while another factor that influences resolution is the X‐ray scattering factor of the tissue components. The microscope is designed to give X‐rays of different wavelength by change of target material, and of different penetrating power by altering the voltage. At the time this work started, there was insufficient information available about the conditions required for obtaining useful photographs of longitudinal and cross‐sections of bone, so that the first need was to investigate the effect of variations in the experimental method. These will be considered under three main headings: the instrument, preparation and photography of specimens, target material and voltage. An attempt will also be made to give a preliminary assessment of the areas of investigation where the instrument is likely to prove of greatest use in bone research.
A versatile field‐ion microscope for metallographic studies
Authors: Davies D. M. , Ralph B.
Published: August 2011

The design of a simple and robust field‐ion microscope for making routine metallurgical investigations is presented. Particular attention is given to the choice of ancillary facilities such as the cooling and image‐intensifying systems. Examples of the applications of this microscope are given.
Some observations on the relative dehydration rates of negative stains and biological objects
Authors: Johnson M. W. , Horne R. W.
Published: August 2011

In an attempt to explain the possible mechanism of negative staining a series of experiments are described in which the relative drying rates of two commonly used negative stains and two biological specimens were measured. Turnip yellow mosaic virus and phosphatidyl‐L‐serine were used to represent biological specimens and their dehydration rates were measured against potassium phosphotungstate or ammonium molybdate after tritiated water had been added to the solutions containing protein, lipid, negative stains or mixtures of both. From these experiments it was possible to demonstrate that the protein or lipid retains water for longer periods than the negative‐staining material. The results are in agreement with an earlier proposal that some negative stains set in a rigid ‘glass’ and thus preserve the biological structures before the latter are completely dehydrated either in air on the supporting films, or when finally dried in the vacuum of the electron microscope.