Quantitative electron diffraction evidence for one-dimensional ordering in magnetite above the Verwey transition.

Energy-filtered electron diffraction and three-dimensional reciprocal lattice mapping was used to study the nature of diffuse scattering in magnetite above the Verwey transition temperature. Characteristic Huang scattering associated with a single molecular polaron is observed at room temperature. As the temperature is lowered, the experiment shows narrowing of diffuse scattering in the (001) directions and additional ringlike diffuse scattering at q approximately 0.8, which suggests the presence of one-dimensional structures above the Verwey transition. Experimental measurements of temperature-dependent correlation lengths and diffuse scattering intensity indicate an increase in the number and length of the one-dimensional structure as the temperature is cooled toward the transition. This study demonstrates the electron sensitivity to atomic displacement and the quality of electron diffraction data for studying phase transition in complex materials.

Use of buserelin to induce ovulation in the cyclic mare.

Inducing ovulation in a cyclic mare is often necessary. For this purpose, hCG has been used commonly, but the response can be reduced after successive administrations. The aims of this study were to test the effectiveness of buserelin in hastening ovulation in estrus mares, and its influence on fertility; and to investigate the effect of treatment on LH secretion. Five crossover trials were designed to compare the effect of two treatments: buserelin (40 microg in 4 doses i.v. at 12 h intervals) vs placebo (Experiments 1 and 2); buserelin 40 microg (in 4 doses i.v.) vs 20 microg (Experiment 3); buserelin (4 doses of 20 microg i.v.) vs hCG (1 dose of 2,500 IU i.v.) (Experiment 4); or buserelin (3 doses of 13.3 microg at 6 h interval) vs hCG (Experiment 5). In Experiment 2, blood samples were taken hourly until ovulation, for LH measurements. In Experiment 1, buserelin treatment significantly hastened ovulation. Reduction of the dose by half (Experiment 3) did not alter the effectiveness. In Experiments 4 and 5, buserelin was as effective as hCG in inducing ovulation between 24 and 48 h after initiation of treatment. Buserelin treatment induced a rise in LH concentration during the 48 h period of the experiment, and LH concentrations before ovulation were significantly higher in buserelin treated cycles than in placebo cycles. These experiments demonstrated the usefulness of two new protocols of administration of buserelin, as an alternative to hCG for induction of ovulation. One hypothesis explaining the mechanism of action is that the persistant rise in LH concentration could modify the ratio of biological/immunological LH, as it occurs physiologically, thereby hastening ovulation.

Steroid concentrations in follicular fluid aspirated repeatedly from transitional and cyclic mares.

The objectives of the present study were to determine follicular progesterone (P4) and estradiol-17beta (E2) in transitional mares and to compare follicular steroid concentrations between transitional and cyclic mares. Follicles > 8 mm were aspirated under transvaginal ultrasound-guidance 4 times at 3 to 4 day intervals (T1-T4) in Norwegian pony mares during vernal transition. During the breeding season, follicular aspirations were conducted in each mare on Day 6, Day 14 and Day 18 after ovulation of 3 separate estrous cycles (Day of ovulation = Day 0). Plasma and follicular fluids were analyzed for P4 and E2 with ELISA and RIA, respectively. Plasma P4 concentrations remained below 1 ng/mL throughout T1-T4, while the follicular P4 concentrations increased significantly to cyclic levels after the first transitional aspiration. Plasma E2 concentrations similarly remained at low levels during the course of the transitional aspirations, while the follicular E2 concentrations increased gradually over the 4 aspirations to cyclic concentrations. The mares ovulated on average 9.8 +/- 1.6 (mean +/- SEM) days after the last transitional aspiration, and 16.6 +/- 0.2, 11.3 +/- 1.5 and 23.2 +/- 4.4 days after aspirations conducted on Day 6, 14 and 18, respectively. The present study demonstrates that in the transitional mare newly developing follicles exhibit biosynthesis of P4 and E2. Furthermore, an increase in follicular steroid concentrations is not necessarily reflected in the peripheral steroid concentrations.

Experimental measurement of electron diffuse scattering in magnetite using energy-filter and imaging plates

We have measured the diffuse scattering in magnetite as a function of temperature using the LEO 912Ohms energy-filtering electron microscope and the imaging plates. This study takes the advantage of the Koehler illumination system, energy filtering and the imaging plates for recording electron diffraction pattern over a large dynamic range. The experiment clearly shows a quantitative change in diffuse scattering distribution, which has the characteristics of one-dimensional ordering. This study clearly demonstrates the possibility for the quantitative study of diffuse scattering using electron diffraction.

Use of quantitative convergent-beam electron diffraction in materials science.

Methods for quantitative convergent-beam electron diffraction are outlined and some results of our applications of convergent-beam electron diffraction are shown, with emphasis on quantitative analysis of crystal structures in materials science. Examples of thickness measurements and determination of lattice parameters are presented. Measurements of low-order structure factors to obtain information on bonding charge-density distributions are reviewed, with examples from TiAl intermetallics. For non-centrosymmetric crystals, a method to determine three-phase structure invariants is given. Determination of polarity is also discussed.

Three-phase structure invariants and structure factors determined with the quantitative convergent-beam electron diffraction method.

Quantitative convergent-beam electron diffraction is used to determine structure factors and three-phase structure invariants. The refinements are based on centre-disc intensities only. An algorithm for param-eter-sensitive pixel sampling of experimental intensities is implemented in the refinement procedure to increase sensitivity and computer speed. Typical three-beam effects are illustrated for the centrosymmetric case. The modified refinement method is applied to determine amplitudes and three-phase structure invariants in noncentrosymmetric InP. The accuracy of the results is shown to depend on the choice of the initial parameters in the refinement. Even unrealistic starting assumptions and incorrect temperature factor lead to stable results for the structure invariant. The examples show that the accuracy varies from 1 to 10 degrees in the electron three-phase invariants determined and from 0.5 to 5% for the amplitudes. Individual phases could not be determined in the present case owing to spatial intensity correlations between phase-sensitive pixels. However, for the three-phase structure invariant, stable solutions were found.

Development and validation of an improved enzyme-linked immunosorbent assay for the detection of thyroglobulin autoantibodies in canine serum samples.

An enzyme-linked immunosorbent assay to detect thyroglobulin autoantibodies (TGAB) in canine serum was developed and validated. The test result for each sample was derived from the optical density readings (OD) and expressed as an Ab-score(%) calculated from three in-house calibrators. The assay specifically detected TGAB as judged from lack of response in the assay after samples had been incubated with specific antigen. Intra- and interassay coefficients of variation ranged from 2.0-4.9% and 4.6-9.9%, respectively. The detection limit, an Ab-score of 5.6%, was close to the median Ab-score of 10% observed in healthy dogs (n = 132). The median Ab-score of dogs with primary hypothyroidism and lymphocytic thyroiditis (n = 11), skin diseases (n = 35), and non-thyroidal diseases (n = 63) was 340%, 12%, and 8%, respectively. The prevalence of TGAB in hypothyroid dogs with lymphocytic thyroiditis (sensitivity) was 91% (95% confidence limits: 59%-99%). In dogs with dermatological diseases without lymphocytic thyroiditis the prevalence of TGAB was 3% corresponding to a specificity of 97% (95% confidence limit: 85%-100%). In dogs with non-thyroidal diseases and healthy dogs the prevalence of TGAB was 5% and 6%, respectively. The diagnostic accuracy of serum TGAB was evaluated by subjecting the data from 11 dogs with lymphocytic thyroiditis and 35 control dogs without lymphocytic thyroiditis to receiver-operating characteristic curve analysis. The area under the receiver-operating characteristic curve (W = 0.966; 95% confidence limit 87%-100%) was significantly higher than that of a worthless test (0.5) (P < 0.0001), thereby indicating that serum TGAB measurements distinguished between dogs with and without lymphocytic thyroiditis.

Evaluation of the analytical performance of an enzyme immunometric assay (EIA) designed to measure endogenous thyroid-stimulating hormone (TSH) in canine serum samples.

The analytical performance of a commercially available enzyme immunometric assay (EIA) designed to measure endogenous thyroid-stimulating hormone (cTSH) concentration in canine serum samples was evaluated. The precision of the assay was acceptable with intra- and inter-assay coefficients of variation (CV) of 5 and 11%, respectively. The assay maintained linearity under dilution and recovered added cTSH in a linear and proportional manner. The limit of detection (0.03 microgram/L) was well below the values measured in serum from dogs with primary hypothyroidism (median value 1.52 micrograms/L). Further, the assay clearly distinguished dogs with primary hypothyroidism from clinically healthy dogs (P = 0.0004). Based on these findings, it was concluded that the analytical performance of the assay was acceptable.

Establishment of the block against sperm penetration in parthenogenetically activated bovine oocytes matured in vitro.

The ability of a single electric pulse to mimic a block against sperm penetration in bovine oocytes matured in vitro was investigated. Confocal laser scanning microscopy detected a global loss of spots, presumed to be cortical granules, stained with Lens culinaris agglutinin, in pulsed oocytes. Transmission electron microscopy revealed that cortical granule exocytosis occurred within 1 min of stimulation and the number of remaining cortical granules was significantly reduced in all pulsed oocytes. The ability of pulsed oocytes to undergo fertilization in vitro was also affected, as only 31% of the pulsed oocytes were penetrated compared with 87% in the control group. Since incidences of penetration in pulsed oocytes (31%), and of polyspermy in control oocytes (18%) did not differ and were highly correlated (P = 0.009) among trials (n = 15), the induced block is considered to be comparable with the natural block triggered by a spermatozoon. The increased resistance of the zona pellucida to pronase E observed in pulsed oocytes suggests that the induced block depends, at least partly, on modifications of zona pellucida glycoproteins. Finally, the majority (66%) of pulsed, penetrated oocytes did not form male pronuclei, probably as a consequence of asynchrony between the formation of female pronucleus and sperm penetration. The reduced ability of the cytoplasm to induce the formation of a male pronucleus was accompanied by a fall in histone H1 kinase activity to basal values by 3 h after stimulation. These results demonstrate that a single electric pulse can induce a block against sperm penetration similar to that of the spermatozoon itself.

Evaluation of the urinary cortisol: creatinine ratio in the diagnosis of hyperadrenocorticism in dogs.

The diagnostic accuracy of the urinary cortisol:creatinine ratio (CCR), with the cortisol being measured by ELISA, was evaluated by subjecting data from 18 dogs with and 20 dogs without hyperadrenocorticism to receiver operating characteristic (ROC) curve analysis. The area under the ROC curve (W 0.93, SE(w) 0.044) was much higher than 0.5, indicating that the CCR did distinguish between dogs with and without hyperadrenocorticism. A cutoff value of about 60 x 10(-6) was associated with the highest sensitivity (1.0) and specificity (0.85). At the disease prevalence rate of the present study (0.47), the positive and negative predictive values were 0.87 and 1.0, respectively. These numbers indicate that canine hyperadrenocorticism may be safely excluded when the CCR is below 60 x 10(-6) but that a test of higher specificity (eg, the ACTH stimulation test) should be used to confirm the diagnosis of canine hyperadrenocorticism when the CCR is above 60 x 10(-6).

Effect of two virus inactivation methods: electron beam irradiation and binary ethylenimine treatment on determination of reproductive hormones in equine plasma.

Ionizing irradiation and binary ethylenimine treatment have previously been shown to be effective for in-vitro inactivation of virus in biological material. In the present study the 2 methods were tested for possible effects on measurable concentrations of reproductive hormones in equine plasma (luteinizing hormone (LH), folliclestimulating hormone (FSH), progesterone (P4), and oestradiol-17 beta (E2)). The inactivation methods were electron beam irradiation with a dose from 11 to 44 kGy or treatment with binary ethylenimine (BEI) in concentrations of 1 and 5 mmol/L. Generally, there was a close correlation (r > 0.8, < 0.001) between pre- and post-treatment hormone levels. Thus, the different phases of the oestrous cycle could be distinguished on the basis of measured hormone concentrations of treated samples. However, both treatments significantly changed hormone concentrations of the plasma samples. For LH, FSH, and E2 the effect of irradiation and BEI treatment was depressive and dose-dependent. For P4 the effect of irradiation was also depressive and dose-dependent. However, the highest dose of BEI resulted in an increase of measured P4 concentration, which may be attributed to changes in the plasma matrix due to the treatment. Although the treatments affected measured hormone concentrations, the close correlation between pre-treatment and post-treatment measurements means that the diagnostic value will remain unchanged.

Prevalence of antibodies to lipid A in Danish cattle.

A cross-sectional study was performed on the occurrence of IgG antibodies to lipid A of the Gram-negative bacterial lipopolysaccharide (LPS, endotoxin) on serum of 2272 cattle distributed on 19 Danish dairy herds. The relationship between the concentration of antibodies to lipid A (ALI) and age, herd, pregnancy rate and occurrence of mastitis, bovine virus diarrhoea (BVD), reproductive and digestive disorders, diarrhoea, pneumonia, foot disorders, various infections and traumatic udder lesions was investigated. ALI generally was low in calves and increased during their first 1.5 years of life to a steady state, which could be altered by the occurrence of disease. There were significant differences in the mean ALI among the herds (P < 0.001). High ALI was associated with a low herd pregnancy rate, to preceding occurrence of mastitis (P < 0.048), BVD (P < 0.01), reproduction diseases (P < 0.041) and digestion disorders (P < 0.064) in animals older than 2 years. The calf mortality rate was not associated to ALI and there was no correlation between the ALI in calves and their dams. The occurrence of high ALI levels on a herd basis may be an indication of increased challenge or enhanced immunological defense to Gram-negative bacteria or endotoxin.

Evaluation of an immunoradiometric assay for thyrotropin in serum and plasma samples of dogs with primary hypothyroidism.

A commercially available immunoradiometric assay was used to measure the thyrotropin (TSH; thyroid-stimulating hormone) concentration in the serum and plasma of 23 dogs. The basal concentration in five dogs with histologically confirmed primary hypothyroidism (median 0.18 microg/l, range 0.16-0.72 microg/l) was slightly, but not significantly, higher than that in 13 clinically healthy dogs (median 0.09 microg/l, range 0.06-0.34 microg/l). The TSH values in 11 euthyroid dogs with various dermatological diseases (median 0.09 microg/l, range 0.05-0.53 microg/l) were significantly lower than in the hypothyroid dogs, but there was considerable overlap. The assay alone was therefore not capable of giving a firm diagnosis.

Determination of canine prolactin by use of a commercial heterologous enzymeimmunoassay kit.

The present study evaluated a commercial heterologous enzymeimmunoassay (EIA) method for the determination of prolactin (PRL) in canine blood plasma and compared the results found with this heterologous method to the PRL concentrations measured by use of a commercial homologous PRL EIA. The heterologous PRL EIA was characterized by intra-assay coefficients of variation (CV %) of 3-5 % for replicate determinations, and inter-assay CV %'s of 5-7 % between means of duplicate determinations. The least detectable concentration was 0.1 microgram/L. A satisfying accuracy was settled by a recovery of added PRL not different from 100 % at varying PRL levels. The cross-reactivity (CR) to canine growth hormone (GH) was 5 %. Both methods outlined equally well the changes in PRL concentrations induced by administration of metoclopramid and bromocriptin; the specificity of both methods was further confirmed by a lack of response to clonidin GH stimulation test. The results achieved with the two methods were not identical despite of identical dose response relations. This difference in test results was, however, considered to be of minor practical importance if locally established reference ranges are used for evaluation of test results.

Determination of insulin-like growth factor 1 in dogs using a commercially available immunoradiometric assay1).

The objective of the present study was to characterize a commercially available immunoradiometric assay (IRMA) intended for the determination of human insulin-like growth factor 1 (IGF-1) when used to determine IGF-1 in canine plasma samples. The assay was evaluated by assessment of the precision, accuracy, and detection limit, and by comparing the IGF-1 values observed in healthy dogs and in dogs with acromegaly and hereditary pituitary dwarfism to values reported in the literature. Since extraction of IGF-1 from its binding proteins is essential for reliable test results, the efficiency of the extraction method used in the assay was also examined. The main conclusions can be summarized as follows: The commercially available IRMA method offers a precise determination of the IGF-1 concentration in canine plasma samples with IGF-1 values directly comparable to those obtained by others. The assay accuracy was confirmed for normal dogs by parallelity between IGF-1 concentrations and plasma dilutions, whereas plasma from acromegalic dogs could not fullful parallelity. The detection limit was below the IGF-1 concentrations observed in dogs with hereditary pituitary dwarfism. In plasma samples with low or normal IGF-1 levels, the assay's extraction efficiency was comparable to that found using other extraction methods. In samples with IGF-1 concentrations above usual physiological levels, the extraction efficiency decreased unexplainably. However, this decrease in extraction efficiency seems to be of only minor importance when the measurements are used in the diagnosis of canine acromegaly.

An improved radioimmunoassay for the determination of canine growth hormone based on commercially available reagents.

Determination of canine growth hormone (cGH) is of importance when evaluating the growth hormone status in dogs, for instance when diagnosing pituitary dwarfism and acromegaly. At present, analysis for the cGH content in canine blood has mainly been restricted to specialized laboratories which have had the capacity to develop suitable methods. We therefore formulated the objective of the present study to be the development of a generally accessible method utilizing commercially available reagents. We describe a radioimmunoassay (RIA) for the determination of cGH which was based on reagents developed for the determination of porcine growth hormone (pGH), thereby drawing benefit from the similarity between growth hormones from the two species. The RIA method was characterized by satisfying precision and accuracy parameters: intra- and inter-assay coefficients of variation (CV%) were 4-27% and 4-20%, respectively, depending on the cGH concentration of the samples. Recovery of added cGH and parallelism between standard curve and sample dilutions were acceptable, thus indicating identity between the calibrator system and the material under investigation (i.e. plasma samples). When measuring individual plasma samples or samples taken during clonidin response tests from normal dogs, dogs suffering from pituitary dwarfism and from dogs suffering from acromegaly, the results were comparable with previous findings.

Evaluation of an enzyme linked immunosorbent assay for the determination of free thyroxine in canine plasma samples assisted by data on biological variation.

The purpose of the present study was to evaluate a commercially available enzyme linked immunosorbent assay (ELISA) for the determination of the free thyroxine (FT4) concentration in canine plasma samples. The test kit Enzymun-Test (Boehringer Mannheim) for determination of FT4 concentrations in human serum and plasma was used in connection with the analyzer Enzymun-Test(Boehringer Mannheim) System ES-33 by following the assay protocol suggested by the manufacturer. To assist in the evaluation of the ELISA method, goals for the precision of the assay were established from the inter- and intraindividual coefficients of variation estimated from determination of the plasma FT4 concentration in blood samples collected weekly from 8 healthy dogs for a period of 4 weeks. The intra- and interassay coefficients of variation ranged from 6% to 13%, the proposed goals for the precision ranging between 10% and 15%. The detection limit of the assay was 2 pmol/l which is well below the FT4 concentration commonly observed in plasma from dogs, and the FT4 concentrations determined in plasma from healthy dogs were comparable to those reported in the literature. Hence, the ELISA method evaluated in this study offers a precise and accurate measurement of the FT4 concentration in canine plasma samples, the inter- and intraassay precision comparing favorably to the proposed specifications for assay precision using estimates of the inter- and intraindividual coefficients of variation.

Endocrine correlates of placental permeability during gestation in the pig.

1. Hormone and electrolyte concentrations were determined in amniotic and allantoic fluid collected from sows at 47-112 days of gestation and correlated with placental and fetal membrane electrolyte permeability. 2. Significant differences between the electrolyte and endocrine gestational profiles of amniotic and allantoic fluid were identified. 3. The principal endocrine correlate (r > 0.62; P < 0.01) of gestational changes in the sodium and chloride permeabilities of the porcine placenta and fetal membranes was found to be the concentration of oestrogen in the amniotic fluid.

Clinical chemical diagnosis of diseases assisted by logistic regression illustrated by diagnosis of canine primary and secondary hepatobiliary diseases.

OBJECTIVE: The purpose of the present study was to demonstrate the use of logistic regression models in the prediction of diseases using the prediction of canine primary and secondary hepatobiliary diseases as an example. Briefly, in a logistic regression model independent variables (i.e. the analytical results) are combined in a linear equation that is used to estimate the logarithm of the odds (logit) of an event (i.e. having primary or secondary hepatobiliary disease). From the estimated logit given by the logistic regression model, a conditional probability of the event (i.e. having primary or secondary hepatobiliary disease) can be calculated. STUDY DESIGN: Twenty-six dogs with verified primary and secondary hepatobiliary diseases and 19 dogs, initially suspected to have hepatobiliary diseases, but with apparently other diseases, were included in the study. The following clinical chemical parameters were measured: alanine aminotransferase (ALAT), aspartate aminotransferase (ASAT), alkaline phosphatase (AP), bilirubin(Total) (TB), urea, glucose, retention of bromosulphthalein (BSP), fasting and postprandial total serum bile acid concentration (FSBA and PSBA). Logistic regression analysis, using the CATMOD procedure in SAS, was used to select which of the measured parameters should be included in the model, and to derive a logistic regression model using the selected parameters. To observe more closely the potential of the logistic regression model, the model was also used to classify a test group consisting of 13 dogs (6 dogs with hepatobiliary diseases and 7 dogs with other diseases). RESULTS: By logistic regression analysis, ASAT and PSBA were selected to be included in the final model, and the final logistic regression model was Y = -3.194 + 0.044.PSBA + 3.251.ASAT. The logistic regression model classified correctly 38 (84%) of 45 dogs in the present study. Specifically, 21 (81%) of 26 dogs with verified primary or secondary hepatobiliary diseases and 17 (90%) of 19 dogs with various other diseases were correctly classified by the logistic regression model. When the model was used on the test group, 5 (83%) of 6 dogs with hepatobiliary diseases and 7 dogs (100%) of 7 dogs with other diseases were correctly classified. CONCLUSIONS: Even though the logistic regression model derived in the present study only serves as an example, thus reducing the practical usefulness of the derived logistic regression model, the present study indicates a great potential of logistic models for the diagnosis of diseases.