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Structure of the nucleoid in cells of Streptococcus faecalis.

The structure of the nucleoid of Streptococcus faecalis (ATCC 9790) was examined and compared in the unfixed and fixed states by immersive refractometry and electron microscopy. It appears from these studies that the nucleoid structure is much more centralized in unfixed chloramphenicol-treated (stationary-phase) cells than it is in cells in the exponential phase of growth. The more dispersed configuration of the exponential-phase nucleoid could be preserved by fixation in glutaraldehyde, but not in Formalin or in osmium tetroxide. One important factor in explaining these differences in preservation is that glutaraldehyde (but not Formalin or osmium tetroxide) can rapidly cross-link the amino groups of macromolecules in cells. It was also observed that osmium tetroxide resulted in a preferential breakdown of nascent ribonucleic acid. These results are interpreted as indicating that glutaraldehyde is able to stabilize the exponential-phase nucleoid before it assumes the more central appearance seen in osmium tetroxide- and Formalin-fixed cells. These results are discussed in terms of the proposed organization of the exponential-phase nucleoid in unfixed cells.

Cell Nucleus↗

Freeze-drying of cephalothin sodium: granularly agglomerated crystallization during freezing. II.

The mechanism of granularly agglomerated crystallization during freezing of cephalothin sodium (CET-Na) in aqueous solution has been discussed. Our previous report noted that strict control of the following is important in order to obtain crystalline granular agglomerates of CET-Na: (1) thermal history of the solution before receiving freezing, (2) cooling rate in freezing, and (3) aging temperature level in the crystal growth step. In order to clarify the physico/chemical meanings of the individual controls, further investigations have been made: 1) with varying thermal history, aqueous 30-% CET-Na solutions were prepared for storage, first at 0 degrees C with its supersaturation and secondly at 25 degrees C, in an unsaturated state for observing any structural changes by viscosimetry, refractometry, and surface tensiometry; 2) morphological changes in ice crystals during freezing at varying cooling rates, as well as those during the crystal growth step, were observed by polarized-light cryomicroscopy and scanning electron microscopy; 3) melting, as well as crystal growth, at several aging temperature levels were observed by electrical conductometry and also by the above-mentioned techniques.(ABSTRACT TRUNCATED AT 250 WORDS)

Cephalothin↗

Using light scattering to determine the stoichiometry of protein complexes.

The stoichiometry of a protein complex can be calculated from an accurate measurement of the complex's molecular weight. Multiangle laser light scattering in combination with size-exclusion chromatography and interferometric refractometry provides a powerful means for determining the molecular weights of proteins and protein complexes. In contrast to conventional size-exclusion chromatography and analytical centrifugation, measurements do not rely on the use of molecular weight standards and are not affected by the shape of the proteins. The technique is based on the direct relationship between the amount of light scattered by a protein in solution and the product of its concentration and molecular weight. A typical experimental configuration includes a size-exclusion column to fractionate the sample, a light-scattering detector to measure scattered light, and an interferometric refractometer to measure protein concentration. The determination of the molecular weight of an anthrax toxin complex will be examined to illustrate how multiangle laser light scattering can be used to determine the stoichiometry of protein complexes.

Bacillus anthracis↗

Measurement of urine relative density using refractometer and reagent strips.

The relative density of urine is the ratio of its density to that of water and depends on both the number and weight of solute particles in the sample, while osmolality depends only on the number of solute particles. Water metabolism is regulated by the interaction of the renal medullary countercurrent system with the circulating levels of antidiuretic hormone and thirst. The concentration of solids in urine can be measured by weighing, hydrometry, oscillations of a capillary tube, refractometry and reagent strip. These techniques, interrelated but not identical, are commonly used in hospital laboratories and in clinical wards. We compared the results obtained in 1725 urine samples of inpatients and outpatients using an automated refractometer to those obtained using two visually read dip stick tests. The correlation coefficients (Super Aution analyser vs. Aution Sticks 10EA, Aution Sticks 10 EA vs. N-Multistix, Super Aution analyser vs. N-Multisticks were 0.663, 0.645 and 0.514, respectively) and the great dispersion of mountain plots demonstrates that different techniques are not interchangeable in the measurement of relative density. Since the results obtained after discarding the samples with pH higher than 7 and those containing glucose or protein were very similar to the ones reported above, the role of these interferents appears negligible in inducing the discrepancy.

Humans↗

Passive transfer of colostral immunoglobulins in calves.

Passive transfer of colostral immunoglobulins has long been accepted as imperative to optimal calf health. Many factors, including timing of colostrum ingestion, the method and volume of colostrum administration, the immunoglobulin concentration of the colostrum ingested, and the age of the dam have been implicated in affecting the optimization of absorption. The practice of colostrum pooling, the breed and presence of the dam, and the presence of respiratory acidosis in the calf also may affect passive transfer. Various tests have been reported to accurately measure passive transfer status in neonatal calves. The radial immunodiffusion and the enzyme-linked immunosorbent assay (ELISA) are the only tests that directly measure serum IgG concentration. All other available tests including serum total solids by refractometry, sodium sulfite turbidity test, zinc sulfate turbidity test, serum gamma-glutamyl transferase activity, and whole blood glutaraldehyde gelation estimate serum IgG concentration based on concentration of total globulins or other proteins whose passive transfer is statistically associated with that of IgG. This paper presents a comprehensive review of the literature of passive transfer in calves including factors that affect passive transfer status, testing modalities, effects of failure of passive transfer on baseline mortality, consequences of failure of passive transfer, and some treatment options. Many previously accepted truisms regarding passive transfer in calves should be rejected based on the results of recent research.

Acidosis↗

Comparison of in vitro tests for evaluation of passive transfer of immunoglobulins in giraffe (Giraffa camelopardalis).

Serum samples from captive giraffe (Giraffa camelopardalis) were tested to assess passive transfer of immunoglobulins using in vitro methods developed for domestic ruminants. Estimated immunoglobulin levels were compared using five tests (protein electrophoresis, total protein refractometry, zinc sulfate turbidity, glutaraldehyde coagulation, and sodium sulfite turbidity). A linear relationship was observed among total protein, gamma globulin (electrophoretic measurement), and immunoglobulin level based on spectrophotometric measurement of zinc sulfate turbidity. Nonquantitative assays also demonstrated statistical correlation with the quantitative methods. Using criteria similar to those established for domestic species, cutoff values for failure of passive transfer (FPT) were established for these tests in neonatal giraffe: 1) total protein <6.0 g/dl; 2) gamma globulin < 0.5 g/dl; 3) estimated immunoglobulin level < 1,000 mg/dl (zinc sulfate turbidity); 4) glutaraldehyde coagulation test negative; or 5) no visually detectable turbidity in 16% sodium sulfite or Bova-S negative. Retrospective examination of the medical histories showed a strong statistical association between animals designated as having FPT and those that were removed from their dams based on clinical assessment to be hand-reared. Application of these tests in the field should allow earlier detection and intervention for FPT in neonatal giraffe.

Animals↗

[Oxidase-peroxidase method for determination of ethanol in fermented musts and wine products].

A new alcohol oxidase-peroxidase method of determination of ethanol content in fermented musts and wine products is described and compared to conventional methods routinely used in winemaking. The sensitivity, accuracy, and reliability of this method were determined. The results of ethanol determination in fermented musts and wines correlated well with the data obtained by refractometry (correlation coefficient R = 0.9595, p < 0.0001) and densitometry (correlation coefficient R = 0.9384, p < 0.0001). This method is less time- and labor-consuming and allows simultaneous testing a series of wine samples.

Alcohol Oxidoreductases↗

A comparison of serum harvesting methods and type of refractometer for determining total solids to estimate failure of passive transfer in calves.

Monitoring the passive transfer of immunoglobulins from colostrums particularly in noncentrifuged samples can be useful for calf health management programs. The results of total solids refractometry from centrifuge and noncentrifuge harvested sources of serum were highly correlated (R2 = 0.95). Results from a digital and a hand-held refractometer were highly correlated (R2 = 0.96).

Animals↗

The clinical refractometer: a useful tool for the determination of specific gravity and osmolality in canine urine.

For the determination of specific gravity of canine urine the urinometer is often used. An alternative method is based on refractometry. Both methods were compared and a table for converting refractometer values to urinometer values is given. The refractometer was also compared with an osmometer. A linear relation was found between the readings of both instruments. The results clearly indicate that the refractometric method is very suitable for the small animal practitioner. The instrument appears to be a good alternative for both the urinometer and the osmometer.

Animals↗

A novel method of measuring the concentration of anaesthetic vapours using a dew-point hygrometer.

The Antoine equation relates the saturated vapour pressure of a volatile substance, such as an anaesthetic agent, to the temperature. The measurement of the 'dew-point' of a dry gas mixture containing a volatile anaesthetic agent by a dew-point hygrometer permits the determination of the partial pressure of the anaesthetic agent. The accuracy of this technique is limited only by the accuracy of the Antoine coefficients and of the temperature measurement. Comparing measurements by the dew-point method with measurements by refractometry showed systematic discrepancies up to 0.2% and random discrepancies with SDS up to 0.07% concentration in the 1% to 5% range for three volatile anaesthetics. The systematic discrepancies may be due to errors in available data for the vapour pressures and/or the refractive indices of the anaesthetics.

Anesthesia, Inhalation↗

[Modification of the lipid membrane matrix with magnesium].

The influence of sulphuric magnesium on the molecular regulation processes in lecithin-water systems was investigated by the optical methods of polarized microscopy, refractometry and infrared spectroscopy. The obtained changes of structure parameters are interpreted by the modification role of Mg2+ cations and SO4(2-) groups.

Magnesium Sulfate↗

Effect of low-dose latrunculin B on anterior segment physiologic features in the monkey eye.

OBJECTIVES: To determine if low doses of topical latrunculin B (LAT-B) will increase outflow facility and decrease intraocular pressure without damaging the cornea and if they will inhibit miotic and accommodative responses to pilocarpine in monkeys. METHODS: We measured intraocular pressure (Goldmann tonometry) before and after 1 and 9 doses of 0.005% and 0.01% topical LAT-B and vehicle given twice daily on successive weeks; outflow facility (perfusion) following 15 doses; central corneal thickness (ultrasonic pachymetry) before and after 1 and 9 doses of 0.01% LAT-B and vehicle; pupillary diameter (calipers); and accommodation (refractometry) before and after 1 dose of 0.005% and 0.02% LAT-B. RESULTS: Latrunculin-B dose-dependently decreased intraocular pressure, multiple doses more than a single dose. Maximal mean +/- SEM hypotension after 1 dose was 2.5 +/- 0.3 mm Hg (0.005% LAT-B; n = 8; P<.001) or 2.7 +/- 0.6 mm Hg (0.01% LAT-B; n = 8; P<.005); maximal mean +/- SEM hypotension after 9 doses was 3.2 +/- 0.5 mm Hg (0.005% LAT-B; n = 8; P<.001) or 4.4 +/- 0.6 mm Hg (0.01% LAT-B; n = 8; P<.001). Outflow facility was increased by mean +/- SEM 75% +/- 13% (n = 7; P<.005). Central corneal thickness was not changed after 1 or 9 doses of 0.01% LAT-B. Miotic and accommodative responses to intramuscular pilocarpine were dose-dependently inhibited. With 0.02% LAT-B, inhibition of miosis was substantial, whereas the inhibition of accommodation was only about 25%. With 0.005% LAT-B, the effects were trivial. CONCLUSIONS: In ocular normotensive monkeys, 0.005% and 0.01% LAT-B administered topically increases outflow facility and/or decreases intraocular pressure without corneal effects. Multiple doses reduce intraocular pressure more than a single dose. Latrunculin-B dose-dependently relaxes the iris sphincter and ciliary muscle, with some separation of miotic and accommodative effects. Clinical Relevance Multiple treatments with low topical doses of LAT-B may substantially reduce outflow resistance in eyes with glaucoma without adversely affecting the cornea.

Accommodation, Ocular↗

A human model of allergic conjunctivitis.

Using the method of refractometry to measure protein concentration in tears, a simple model was developed to evaluate the allergic response in humans. Timothy grass pollen was instilled into the cul de sac of a human subject, and the protein content of the subject's tears was sampled every 15 minutes for three hours. Once the time course of this experiment was predictable, various eye medications were instilled into the subject's eye 30 minutes after allergic challenge. It was noted that 1.0% and 0.1% prednisolone, each combined with 0.12% phenylephrine hydrochloride, as well as 1% epinephryl borate eye drops, produced an almost immediate return to normal of tear protein levels, whereas 1.0% medrysone, 0.1% fluorometholone, 1.0% prednisolone, and 0.12% prednisolone had less pronounced effects on reduction of elevated tear protein concentration.

Administration, Topical↗

Outflow facility and its response to pilocarpine decline in aging rhesus monkeys.

Refractive error and total outflow facility were determined by Hartinger coincidence refractometry and two-level constant-pressure perfusion, respectively, in 17 rhesus monkeys, aged 5 to 29 years. Maximum accommodative response to corneal (iontophoretic) carbachol hydrochloride, baseline outflow facility, and the facility response to strong but submaximal intracameral doses of pilocarpine hydrochloride all declined with age. The correlation between accommodative response to carbachol and facility response to pilocarpine was slightly stronger than that between age and facility response. Since the ciliary muscle plays a major role in controlling both outflow facility and accommodation, and since histologic and videographic techniques demonstrate an age-related decline in rhesus ciliary muscle excursion induced by topical pilocarpine or electrical stimulation of the Edinger-Westphal nucleus, the present data support the hypothesis that an age-related decline in ciliary muscle mobility is associated, perhaps causally, with an age-related decline in facility and facility responsiveness to cholinergic drugs.

Accommodation, Ocular↗

Amplitudes of accommodation of primate lenses refilled with two types of inflatable endocapsular balloons.

OBJECTIVE: To investigate the maintenance of ocular accommodation by refilling the lenses of the eyes of youthful primates with inflatable endocapsular balloons. METHODS: The lenses of 21 cynomolgus monkeys were refilled following endocapsular phacoemulsification with a balloon that either approximates the shape of the non-accommodated lens or the accommodated crystalline lens. RESULTS: In nine of 15 successfully refilled lenses, we were able to perform automated refractometry to determine the amplitude of accommodation, ie, changes in refraction before and 1 hour after application of topical 4% pilocarpine hydrochloride to alter lens shape. At 2 weeks, 2 to 3 months, and 6 to 12 months after operation, mean (+/- SD) accommodation was 4.6 +/- 2.5, 2.5 +/- 0.5, and 1.7 +/- 0.7 diopters (D), respectively, in the lenses refilled with the nonaccommodation balloon (n = 5; preoperative value, 15.2 +/- 1.3 D), and it was 1.9 +/- 0.5, 1.3 +/- 0.9, and 1.8 +/- 0.9 D, respectively, in the lenses refilled with the accommodation balloon (n = 4; preoperative value, 17.0 +/- 2.9 D). CONCLUSION: The greater yield of accommodation with the nonaccommodation balloon is consistent with the recent theory on the mechanism of accommodation. Although the obtained accommodation was a small fraction of values determined prior to the operation and the small amplitude of accommodation decreased over time, the feasibility of refilling the lens with an inflatable endocapsular balloon, allowing at least some accommodation in the eyes of youthful primates, was demonstrated. Applied to humans, this procedure may allow accommodation following cataract surgery.

Accommodation, Ocular↗

Aging effects on accommodation and outflow facility responses to pilocarpine in humans.

OBJECTIVE: To determine the relationships among age, outflow facility, and refractive and facility responses to pilocarpine in humans. METHODS: Refraction, intraocular pressure, and outflow facility were determined in 30 normal volunteers aged 20 to 75 years, by coincidence refractometry, applanation tonometry, and Schiøtz tonography, respectively, before and 1 hour after a 30-microL drop of 2% or 6% pilocarpine. Simple regression of baseline facility, postpilocarpine facility, and facility change, on age and refractive change singly and jointly, was performed. Stepwise regression models and graphic conditioning plots were used to determine, for each facility variable, its relationship to age or refractive change specifically. RESULTS: Baseline outflow facility and maximum pilocarpine-induced refractive change (ie, accommodation) declined with age, but the decrease in intraocular pressure and the facility response to pilocarpine did not. After adjusting for age, for baseline facility, there was no further relationship to 6% pilocarpine-induced accommodation, and a slight residual relationship to 2% pilocarpine-induced accommodation. After adjusting for both 2% or 6% pilocarpine-induced accommodation, the relationship to age was still significant. The facility increase after 2% or 6% pilocarpine did not depend on age and/or accommodative amplitude. CONCLUSIONS: In humans, as previously described in rhesus monkeys, an age-related loss of ciliary muscle mobility may compromise the basal function of the trabecular meshwork. However, unlike monkeys, humans exhibit no loss of the intraocular pressure or outflow facility response to pilocarpine with age.

Accommodation, Ocular↗

Online monitoring of preferential crystallization of enantiomers.

Polarimetry is used for continuous online monitoring of optical resolution by preferential crystallization. In combination with refractometry the liquid phase composition is determined, allowing one to follow the resolution progress quantitatively. The measurement techniques were calibrated up to relatively high solution concentrations and combined with the crystallizer. The resolution of DL-threonine was performed by preferential crystallization experiments in aqueous solution varying several process parameters like supersaturation, seed amount, initial enantiomeric excess, and scale. The resolution progress can be conveniently described by profiles of the optical rotation (polarimetric signal) and the crystallization pathway in the corresponding ternary phase diagram. The method outlined is applicable for dynamic process optimization and control purposes in "quasi-continuous" chiral separation processes.

Calibration↗

Direct and indirect methods for molar-mass analysis of fragments of the capsular polysaccharide of Haemophilus influenzae type b.

Two methods are described for direct molar-mass measurement of low-molar-mass fragments obtained by oxidative cleavage of the capsular polysaccharide of Haemophilus influenzae type b. Absolute molar masses were determined by size-exclusion chromatography (SEC) with detection by multiangle laser-light-scattering photometry (MALLS) and differential refractometry (RI). The end-group structure of the polysaccharide fragments allowed the direct measurement of average chain length by quantitative 1H NMR, from which molar masses were derived. Variation between the molar masses obtained by the two methods ranged from 5 to 7%. When molar masses estimated by indirect methods were compared to SEC-MALLS/RI data, significant deviations were observed. Analysis by SEC with secondary calibration with dextran standards gave molar masses that exceeded the SEC-MALLS/RI data by as much as 2.5-fold. Molar masses estimated by a combination of colorimetric assays varied from the SEC-MALLS/RI data by as much as 50%. These results demonstrated the applicability and superior accuracy of the direct methods of molar-mass determination of the polysaccharide fragments.

Bacterial Capsules↗