Permanent Turbidity-Standards

Permanent turbidity reference standards suitable for measurement of microbial suspensions were prepared by suspending finely divided titanium dioxide in aryl sulfonamide-formaldehyde or methylstyrene resins. Turbidities of these standards, adjusted to a useful range for microbiological and immunological studies, were compared with other reference standards in use today. Tube holders for a Coleman Photonephelometer and a Nepho-Colorimeter were modified to eliminate the water well and to allow use of optically standardized 10-, 16-, or 18-mm test tubes. The standards and the tube holders have been used satisfactorily for more than 12 years.     

Specimen Holders for Simultaneous Critical Point Drying of Multiple Biological Specimens

Specimen holders were developed for simultaneous critical point drying of multiple specimens of monolayer all cultures grown on Leighton tube glass cover slips or for specimens of ells collected on silver or cellulose membrane filters. The use of these multiple specimen holders makes it possible to process several specimens in parallel and thus significantly reduce the time required for handling large numbers of samples and also eliminate the possible variations that occur in processing individual samples one at a time in series.     

International reference standards in coagulation

Measurement of coagulation factor activity using absolute physico-chemical techniques is not possible and estimation therefore relies on comparative bioassay relative to a reference standard with a known or assigned potency. However the inherent variability of locally prepared and calibrated reference standards can give rise to poor agreement between laboratories and methods. Harmonisation of measurement between laboratories at the international level relies on the availability of a common source of calibration for local reference standards and this is provided by the World Health Organization (WHO) International Standards which define the International Unit for the analyte. This article describes the principles, practices and problems of biological standardisation and the development and use of reference standards for assays of coagulation factors, with particular emphasis on WHO International Standards for both concentrates and plasma.     

Mechanical study of potential ceramic implant materials for minimal invasive anterior lumbar interbody fusion]

While autologous bone grafts are highly suitable for use in spinal arthrodesis, their use is also associated with problems (traumatization, complications). Ceramic bone substitute materials provide an attractive alternative for lumbar interbody spinal fusion. The aim of the present study was to investigate the mechanical properties of various types of ceramic using a specific fusion method. Ten specimens each of 7 different types of ceramic were tested using a hydraulic testing machine with two different sample holders: polyurethane foam (mechanical properties similar to cancellous bone) and aluminium. The parameters axial compression and axial torque were investigated. With the polyurethane foam holders, none of the ceramic implants failed under compression, while under axial rotation, two types of ceramic failed. With the aluminium holders, 3 ceramics showed no failure up to 25 kN under compression, while under torsion all the ceramics failed. One type of ceramic showed specific fracture properties with a higher load-bearing capacity after failure in comparison with all the other types studied.     

IMPORTANCE OF REFERENCE STANDARDS IN TRAINING PANELISTS

When training a sensory evaluation panel, reference standards play an important role in developing appropriate terminology, establishing intensity ranges and showing the action of an ingredient. In addition, use of reference standards reduces the amount of training time while providing documentation for terminology. Product characteristics can be demonstrated through reference standards for application in plant quality assurance programs as well as for project planning in new product development, product maintenance (i.e., shelf-life), product improvement and cost reduction programs.     

Device for holding a variety of tissue culture vessels during microscopy.

A device for holding tissue culture tubes, plates and flasks has been designed for use in microscopy of cell cultures, which eliminates changing holders when using varied types or sizes of vessel.     

Device for holding a variety of tissue culture vessels during microscopy

Summary A device for holding tissue culture tubes, plates and flasks has been designed for use in microscopy of cell cultures, which eliminates changing holders when using varied types or sizes of vessel.     

Canadian-Trained Basic Medical Scientists, 1946-47—1963-64

The number of master''s and doctoral degree holders who obtained their degree in a basic medical science under the supervision of a Canadian medical faculty between 1946-47 and 1963-64 was obtained from the medical schools. Of the total degree holders, 69% are currently residing in Canada, 23% in the U.S.A., and the remaining 8% in overseas countries.Questionnaire returns from doctoral degree holders revealed that citizenship status at the time of graduation is positively related to migration; migration rates were lowest for Canadian-born and highest for landed immigrants and foreign students. Geographic mobility during training was also found to be a significant factor which increased the propensity to migrate. One-half of those who took further postdoctorate training in the United States are currently living in the United States, compared to 15% of those who received all their training in Canada. Information on current type of employment revealed that only a quarter of the Ph.D. respondents are in a basic science teaching position in Canada.     

Catch-up growth in short-at-birth NICU graduates

The statural catch-up growth, defined as reaching at least tenth length/height percentile (P10) for normal population standards (-1.28 SD score, SDS), was studied in 73 infants short at birth (length < P10 for gestational age) admitted to NICU. Mean gestational age at birth was 35.2 weeks (range 29-41) and mean birth length standard deviation score -2.31 (-4.52/-1.46). Infants were measured at birth, at 3, 6, 12, 18, and 24 months corrected age and then once a year until 6 years chronological age. Statural catch-up growth was studied, with reference both to normal population standards and to individual genetic target. With reference to normal population standards, 44% of infants had caught-up at 3 months of age, 51% at 3 years, 66% at 4 years and 73% at 6 years. In the case of individual genetic targets, a similar trend was present, but the absolute values were slightly higher from 4 to 6 years (73 vs. 66% and 78 vs. 73%, respectively). Statistically significant changes in mean standard deviations score for chronological age were present from birth to 3 months, 3 to 12 months, 3 to 4 years and 5 to 6 years (p<0.05). No differences were found in this trend of recovery when considering ponderal index (PI) at birth (symmetrical vs. asymmetrical), sex (male vs. female) or gestational age (p>0.05). In the majority of cases infants with short stature at birth admitted to a NICU had a statural catch-up growth within the first years of life. This is more evident when considered in relation to individual genetic target rather than to normal population standards.     

Assessing the use of known mutagens to calibrate the Salmonella typhimurium mutagenicity assay: I. Without exogenous activation

There has been an increasing need in genetic toxicology to progress from strictly qualitative tests to more quantitative tests. This, in turn, has increased the need to develop better quality assurance and comparative bioassay methods. In this paper, two laboratories tested 10 Salmonella mutagens in order to determine the usefulness of selected chemicals as potential reference materials to calibrate the Salmonella assay. If variance within a bioassay is sufficiently low and the rankings of the compounds are of acceptable consistency, the chemicals later could be evaluated for use as standard control compounds, as audit materials, and as standard reference materials for comparative bioassay efforts. The results demonstrated that the chosen chemicals (with the possible exception of dimethylcarbamylchloride) provide such consistent results in the Salmonella mutagenicity bioassay that they can be used for semi-quantitative calibration and as possible bioassay controls, special audit chemicals, and potentially as reference standards in comparative bioassay efforts. Reference standards, whether used as audit materials or in comparative bioassays, must be used concurrently with the test substances of interest; used without bias; used in a standardized, highly controlled bioassay; and be tested across an appropriate dose range. The study also shows that when these compounds are used as reference standards much care must be given to the number and spacing of doses if highly reproducible slope values are to be generated. We recommend use of a pilot test to establish a dose range for definitive tests and the placement of doses for the definitive tests within the first half of the linear dose-response curve. For appropriate comparisons, one should replicate the tests using the defined dose range and analyze the results in a non-biased statistical manner.     

18O-labeled proteome reference as global internal standards for targeted quantification by selected reaction monitoring-mass spectrometry

Selected reaction monitoring (SRM)-MS is an emerging technology for high throughput targeted protein quantification and verification in biomarker discovery studies; however, the cost associated with the application of stable isotope-labeled synthetic peptides as internal standards can be prohibitive for screening a large number of candidate proteins as often required in the preverification phase of discovery studies. Herein we present a proof of concept study using an (18)O-labeled proteome reference as global internal standards (GIS) for SRM-based relative quantification. The (18)O-labeled proteome reference (or GIS) can be readily prepared and contains a heavy isotope ((18)O)-labeled internal standard for every possible tryptic peptide. Our results showed that the percentage of heavy isotope ((18)O) incorporation applying an improved protocol was >99.5% for most peptides investigated. The accuracy, reproducibility, and linear dynamic range of quantification were further assessed based on known ratios of standard proteins spiked into the labeled mouse plasma reference. Reliable quantification was observed with high reproducibility (i.e. coefficient of variance <10%) for analyte concentrations that were set at 100-fold higher or lower than those of the GIS based on the light ((16)O)/heavy ((18)O) peak area ratios. The utility of (18)O-labeled GIS was further illustrated by accurate relative quantification of 45 major human plasma proteins. Moreover, quantification of the concentrations of C-reactive protein and prostate-specific antigen was illustrated by coupling the GIS with standard additions of purified protein standards. Collectively, our results demonstrated that the use of (18)O-labeled proteome reference as GIS provides a convenient, low cost, and effective strategy for relative quantification of a large number of candidate proteins in biological or clinical samples using SRM.     

Cross-platform comparison of nucleic acid hybridization: Toward quantitative reference standards

Measuring interactions between biological molecules is vitally important to both basic and applied research as well as development of pharmaceuticals. Although a wide and growing range of techniques is available to measure various kinetic and thermodynamic properties of interacting biomolecules, it can be difficult to compare data across techniques of different laboratories and personnel or even across different instruments using the same technique. Here we evaluate relevant biological interactions based on complementary DNA and RNA oligonucleotides that could be used as reference standards for many experimental systems. We measured thermodynamics of duplex formation using isothermal titration calorimetry, differential scanning calorimetry, and ultraviolet–visible (UV–vis) monitored denaturation/renaturation. These standards can be used to validate results, compare data from disparate techniques, act as a teaching tool for laboratory classes, or potentially to calibrate instruments. The RNA and DNA standards have many attractive features, including low cost, high purity, easily measurable concentrations, and minimal handling concerns, making them ideal for use as a reference material.     

Bacterial endospore inactivation caused by outgassing of vapourous hydrogen peroxide from polymethyl methacrylate (Plexiglas)

AIMS: To investigate the cause and to eliminate the inactivation of Bacillus anthracis strain Sterne spores settled onto agar and stainless steel surfaces in plastic holders. METHODS AND RESULTS: In an experimental chamber in which spores settled onto sampling surfaces, vapourous hydrogen peroxide (VHP) was used for decontamination between experiments. It was demonstrated that hydrogen peroxide (H(2)O(2)) absorbed into plastic (Plexiglas) surfaces and could outgas in the sample holders. Further experiments demonstrated that H(2)O(2) was released from Plexiglas sample holders in sufficient quantity to inactivate spores. High temperature degassing (30-35 degrees C) for several days or aluminum coating of the surfaces were two remedies found to be effective in preventing inadvertent spore inactivation. CONCLUSIONS: H(2)O(2) can be absorbed into plastic and released after an extended period of time (weeks), allowing a sufficient concentration to accumulate in small volumes to inactivate spores. Outgassing the plastic or coating the surface with an impermeable layer are potential solutions to reduce spore inactivation. SIGNIFICANCE AND IMPACT OF THE STUDY: Many studies with bacilli and other organisms are carried out using small plastic containers that may have been sterilized using H(2)O(2) or other agents. This study presents a cautionary note to ensure elimination of H(2)O(2) or other sterilizing agents to prevent spurious results.     

Reference Values for Serum Magnesium Levels in Young Adult Iranian Subjects

This study aims at determining the reference values for serum magnesium (Mg) concentrations in Iranian adults. Serum Mg level was measured using flame atomic absorption spectrometry in 491 subjects (233 men and 258 women), aged 20-50 years, randomly selected from a population-based study. The International Federation of Clinical Chemistry guidelines and the robust method were used for determining the reference values. The 95% reference values for serum Mg concentration were 1.83-2.49, 1.79-2.48, and 1.83-2.55 mg/dL in men, women, and total population, respectively. The prevalences of hypo- and hypermagnesemia, according to the reference values obtained in the current study, were 2.5% and 4.0%, respectively. In conclusion, this study reports serum Mg reference values based on current standards in a large healthy population of young Iranian adults.     

Standardization of high-resolution flow cytometric DNA analysis by the simultaneous use of chicken and trout red blood cells as internal reference standards

Determination of nuclear DNA content by flow cytometry requires comparison with a reference standard. The use of external standards such as lymphocytes or granulocytes is time-consuming and inaccurate. Chicken red blood cells (CRBC) have a DNA content of 35% of the human diploid value and have been widely used as internal standard. The ratio calculated on the basis of the peak channel numbers of the standard and the sample and used to indicate the DNA content (DNA ratio) is, however, very sensitive to changes in the zero level adjustment of the flow cytometer. If two internal standards are used the DNA ratio becomes independent of the zero level. Rainbow trout red blood cells (TRBC) have a DNA content of 80% of human diploid cells. A mixture of CRBC and TRBC was prepared and stored in small aliquots at -80 degrees C. This mixture was added to the sample before staining. The day-to-day variation of the DNA ratio obtained by use of the two standards was smaller than that obtained by CRBC alone. The possibility of sex related differences in DNA content of CRBC and TRBC was examined. The results indicated that a new batch of standards should be tested against the old batch to avoid the introduction of a systematic error.     

Accurate nucleic acid concentrations by nuclear magnetic resonance

Determination of the concentration of biochemical samples often yields values with uncertainties of 10-20% or more. This paper details a protocol for use with 500- to 600-MHz NMR spectrometers to measure approximately 1mM concentrations within +/-1-3% accuracy. With suitable precautions, all compounds have equal NMR "absorption coefficients" for protons. About 2mg of sample are needed for proteins and nucleic acids with MW=5000, although less accurate determinations could be made with smaller amounts. The technique utilizes standardized internal reference reagent compounds, cacodylic acid or 3-(trimethylsilyl)propionic-2,2,3,3-d(4) acid sodium salt. Spectra were signal-averaged using long interpulse delays so that integrals of nonexchangeable protons could be quantified relative to the reference standard. Accurate determinations require careful optimization of the homogeneity of the magnetic field and meticulous attention to sample preparation and spectral processing. The main source of error is usually the accuracy of micropipets. If sample preparation errors could be eliminated, the lower limit of accuracy with the current generation of NMR spectrometers is probably near 0.4%. However, this would require >99.5% sample purity. Methods are described to establish the concentration of the standards, and applications are illustrated with DNA mono- and oligonucleotides. Similar procedures should apply to proteins, polysaccharides, and other biomolecules, with about the same accuracy and precision.     

Social status influences microhabitat selection: breeder and floater Eagle Owls Bubo bubo use different post sites

Social status can be reflected in many aspects of an individual’s behaviour and ecology, including habitat use and conspecific interactions. In territorial species where at least two social groups – breeding birds and non‐territorial floaters – are recognized, the diverse tasks associated with territorial ownership can lead territory holders to behave differently from the non‐territorial part of the population. Territory holders defend their breeding area and reproduce, whereas floating individuals are dispersing and lead a more transient life, during which they do not show any territorial behaviour even when settling in a more or less fixed area (known as the stop phase). As social interactions are based on visual and vocal cues, the use of specific sites for sending and/or receiving signals can be a crucial choice in an animal’s life. By analysing the post‐site selection of Eagle Owl Bubo bubo breeders and floaters during their nocturnal activity, we found that: (1) territory holders selected more visible and dominant posts than non‐territorial floaters; (2) the choice of posts made by floating individuals did not differ between the wandering and stop phases of dispersal; and (3) floating females intruded more frequently than floating males within a breeder’s home‐range. These findings highlight the fact that two social strategies are possible within the same species, depending on an individual’s social status and its related tasks. Breeders could take advantage of visible locations to declare their status as territory holders, whereas floaters could benefit from a more secretive life to wander unnoticed among occupied territories. This secretive life would help floaters to reduce the risks associated with conspecific aggression. Finally, the greater occurrence of floating females within breeders’ home‐ranges can be explained by the fact that female incursions in a breeder’s home‐range are less risky than male intrusions.     

Phytoplankton pigments designation–an application of RP-HPLC in qualitative and quantitative analysis

An account is given of the use of RP-HPLC (reverse-phase high performance liquid chromatography) analysis of high purity chlorophylls and carotenoids standards originating from reference phytoplankton monocultures. All important chromatography parameters were established: retention time, detection limit, spectrum, wavelength of maximum absorption, calibration curve, response factors. Data were also obtained for pigment mixtures in naturalphytoplankton populations in Spitsbergen and the Gulf of Gdask. The identity and purity of all pigments were confirmed by co-elution with standards, including canthaxanthin as an internal standard. The data presented enable the qualitative and quantitative analysis of naturally existing phytoplankton pigments at concentrations down to 0.1 g L^-1 to be carried out without the need for standards from commercial sources.     

Reference standards for determination of DNA content of plant nuclei

Flow cytometry was used to compare 14 potential reference standards for plant DNA content determination. Both chicken and plant internal standards were used, as were propidium iodide (PI) and 4'-6-diamidino-2-phenylindole (DAPI) as fluorochromes. Means and standard errors of the means are presented for the 14 potential reference standards, and the means are compared to those obtained by Feulgen densitometry. Five species are recommended as an initial set of international standards for future plant DNA content determinations: Sorghum bicolor cv. Pioneer 8695 (2C = 1.74 pg), Pisum sativum cv. Minerva Maple (2C = 9.56 pg), Hordeum vulgare cv. Sultan (2C = 11.12 pg), Vicia faba (2C = 26.66 pg), and Allium cepa cv. Ailsa Craig (2C = 33.55 pg). It is recommended that the reference standard of choice be one with 2C and 4C nuclear DNA content peaks similar to, but not overlapping, the 2C and 4C peaks of the target species. We recommend PI as the fluorochrome of choice for flow cytometric determination of plant DNA content. DAPI should be used only if the estimated DNA value is corroborated by using a second stain that has no bias for AT- or GC-rich sequences within genomes.