The subcellular fractionation of embryonic chick tendon and cartilage cells: a re-examination.

A re-examination of the subcellular fractions obtained from matrix-free chick tendon and cartilage cells has been made since the discovery that three out of four of the micrographs of chick tendon microsomal fractions published in an earlier paper from this laboratory were not authentic. The present studies demonstrate that by using the procedures previously reported it is possible to isolate microsomal and submicrosomal fractions from tendon and cartilage cells which exhibit typical morphology when examined by electron microscopy. These observations are consistent with our original biochemical characterization of subcellular fractions, which we know to be valid. Other publications from this laboratory in which these fractionation procedures have been applied to studies of collagen biosynthesis are in no way compromised, and indeed, most of our data have been confirmed by several other laboratories.     

A Model To Estimate the Optimal Sample Size for Microbiological Surveys

Estimating optimal sample size for microbiological surveys is a challenge for laboratory managers. When insufficient sampling is conducted, biased inferences are likely; however, when excessive sampling is conducted valuable laboratory resources are wasted. This report presents a statistical model for the estimation of the sample size appropriate for the accurate identification of the bacterial subtypes of interest in a specimen. This applied model for microbiology laboratory use is based on a Bayesian mode of inference, which combines two inputs: (ii) a prespecified estimate, or prior distribution statement, based on available scientific knowledge and (ii) observed data. The specific inputs for the model are a prior distribution statement of the number of strains per specimen provided by an informed microbiologist and data from a microbiological survey indicating the number of strains per specimen. The model output is an updated probability distribution of strains per specimen, which can be used to estimate the probability of observing all strains present according to the number of colonies that are sampled. In this report two scenarios that illustrate the use of the model to estimate bacterial colony sample size requirements are presented. In the first scenario, bacterial colony sample size is estimated to correctly identify Campylobacter amplified restriction fragment length polymorphism types on broiler carcasses. The second scenario estimates bacterial colony sample size to correctly identify Salmonella enterica serotype Enteritidis phage types in fecal drag swabs from egg-laying poultry flocks. An advantage of the model is that as updated inputs from ongoing surveys are incorporated into the model, increasingly precise sample size estimates are likely to be made.     

A model to estimate the optimal sample size for microbiological surveys

Estimating optimal sample size for microbiological surveys is a challenge for laboratory managers. When insufficient sampling is conducted, biased inferences are likely; however, when excessive sampling is conducted valuable laboratory resources are wasted. This report presents a statistical model for the estimation of the sample size appropriate for the accurate identification of the bacterial subtypes of interest in a specimen. This applied model for microbiology laboratory use is based on a Bayesian mode of inference, which combines two inputs: (ii) a prespecified estimate, or prior distribution statement, based on available scientific knowledge and (ii) observed data. The specific inputs for the model are a prior distribution statement of the number of strains per specimen provided by an informed microbiologist and data from a microbiological survey indicating the number of strains per specimen. The model output is an updated probability distribution of strains per specimen, which can be used to estimate the probability of observing all strains present according to the number of colonies that are sampled. In this report two scenarios that illustrate the use of the model to estimate bacterial colony sample size requirements are presented. In the first scenario, bacterial colony sample size is estimated to correctly identify Campylobacter amplified restriction fragment length polymorphism types on broiler carcasses. The second scenario estimates bacterial colony sample size to correctly identify Salmonella enterica serotype Enteritidis phage types in fecal drag swabs from egg-laying poultry flocks. An advantage of the model is that as updated inputs from ongoing surveys are incorporated into the model, increasingly precise sample size estimates are likely to be made.     

Estimation of gait cycle characteristics by trunk accelerometry

This study reports on the novel use of a portable system to measure gait cycle parameters. Measurements were made by a triaxial accelerometer over the lower trunk during timed walking over a range of self-administered speeds. Signals from each trial were transformed to a horizontal-vertical coordinate system and analyzed by an unbiased autocorrelation procedure to obtain cadence, step length, and measures of gait regularity and symmetry. By curvilinear interpolation, speed-dependent gait parameters could be compared at a normalized speed. It was demonstrated that analysis of gait cycle parameters which previously required fixed laboratory equipment and paced walking procedures, now can be made from data obtained by a timing device and a portable sensor at free walking speeds.     

Towards Harmonisation of Critical Laboratory Result Management - Review of the Literature and Survey of Australasian Practices

Timely release and communication of critical test results may have significant impact on medical decisions and subsequent patient outcomes. Laboratories therefore have an important responsibility and contribution to patient safety. Certification, accreditation and regulatory bodies also require that laboratories follow procedures to ensure patient safety, but there is limited guidance on best practices. In Australasia, no specific requirements exist in this area and critical result reporting practices have been demonstrated to be heterogeneous worldwide.Recognising the need for agreed standards and critical limits, the AACB started a quality initiative to harmonise critical result management throughout Australasia. The first step toward harmonisation is to understand current laboratory practices. Fifty eight Australasian laboratories responded to a survey and 36 laboratories shared their critical limits. Findings from this survey are compared to international practices reviewed in various surveys conducted elsewhere. For the successful operation of a critical result management system, critical tests and critical limits must be defined in collaboration with clinicians. Reporting procedures must include how critical results are identified; who can report and who can receive critical results; what is an acceptable timeframe within which results must be delivered or, if reporting fails, what escalation procedures should follow; what communication channels or systems should be used; what should be recorded and how; and how critical result procedures should be maintained and evaluated to assess impact on outcomes.In this paper we review the literature of current standards and recommendations for critical result management. Key elements of critical result reporting are discussed in view of the findings of various national surveys on existing laboratory practices, including data from our own survey in Australasia. Best practice recommendations are made that laboratories are expected to follow in order to provide high quality and safe service to patients.     

Research in nondomestic species: experiences in reproductive physiology research for conservation of endangered felids

Tremendous strides have been made in recent years to broaden our understanding of reproductive processes in nondomestic felid species and further our capacity to use this basic knowledge to control and manipulate reproduction of endangered cats. Much of that progress has culminated from detailed scientific studies conducted in nontraditional laboratory settings, frequently at collaborating zoological parks but also under more primitive conditions, including in the field. A mobile laboratory approach is described, which incorporates a diverse array of disciplines and research techniques. This approach has been extremely useful, especially for conducting gamete characterization and function studies as well as reproductive surveys, and for facilitating the development of assisted reproductive technology. With continuing advances in assisted reproduction in rare felids, more procedures are being conducted primarily as service-related activities, targeted to increase effectiveness of species propagation and population management. It can be a challenge for both investigators and institutional animal care and use committees (IACUCs) to differentiate these service-based procedures from traditional research studies (that require IACUC oversight). For research with rare cat species, multi-institutional collaboration frequently is necessary to gain access to scientifically meaningful numbers of study subjects. Similarly, for service-based efforts, the ability to perform reproductive procedures across institutions under nonstandard laboratory conditions is critical to applying reproductive sciences for managing and preserving threatened cat populations. Reproductive sciences can most effectively assist population management programs (e.g., Species Survival Plans) in addressing conservation priorities if these research and service-related procedures can be conducted "on the road" at distant national and international locales. This mobile laboratory approach has applications beyond endangered species research, notably for other scientific fields (e.g., studies of hereditary disease in domestic cat models) in which bringing the laboratory to the subject is of value.     

Design and positioning of a reference information model for clinical laboratories

Clinical laboratories produce on a daily basis huge amounts of data, i.e., test results of chemical determinations. The production and the use of these data should be extremely well structured. This and the urge to improve quality was the reason to design a reference information model for clinical laboratories, abbreviated as RILA. A software toolbox facility was used to the design. RILA can be used in any clinical laboratory, but it can also be made laboratory situation-specific. This is done through the implementation of location-specific and detailed information concerning procedures, instruments, capacities, etc. By doing this, the interaction with the laboratories environment has to be defined as well. Certain application areas such as total quality management, workflow analysis and cost analysis can be facilitated through the implemented reference information model. Furthermore, RILA has to be positioned at a proper location with regard to the existing hospital and laboratory automation modules.     

Fine needle aspiration cytology of the breast. A review of 321 cases with statistical evaluation

To assess the accuracy of fine needle aspirations of the breast performed at our institution, all patients undergoing this procedure between the years 1973 and 1982 were evaluated. Correlation was made between the cytologic and histologic diagnoses whenever possible. Correlations were made with mammography results or clinical impressions when biopsies were not obtained. Our results for sensitivity, specificity, predictive value of a positive diagnosis and predictive value of a negative diagnosis were 65.0%, 100%, 100% and 89.6%, respectively. The data indicate that when adequate, well-prepared samples are submitted to the laboratory, accurate cytologic diagnoses can be made. The high specificity and predictive value of a positive result allow for the early diagnosis, treatment and management of breast cancer.     

Training Nonhuman Primates to Cooperate With Scientific Procedures in Applied Biomedical Research

This report provides a brief overview of aspects of training nonhuman primates who have been, and continue to be, used in this laboratory. The research context involves applied behavioral studies in which animals are trained to perform complex operant behavioral sequences, often in their homecage environment. In such studies, animals have freedom to choose whether to engage in appetitively reinforced behavioral tests that employ neither food deprivation nor fluid management. This background of operant conditioning has provided an insight to, and a context for, animal training both as an adjunct to general laboratory management and as a way to expedite scientific procedures. Thus, training has potential implications for both well-being and scientific quality, although it must be considered an adjunct to the provision of socialization with conspecifics in high quality diverse housing systems and not as an alternative to such provision. The importance of discussion and consideration of alternative procedures cannot be overemphasized.     

Training nonhuman primates to cooperate with scientific procedures in applied biomedical research

This report provides a brief overview of aspects of training nonhuman primates who have been, and continue to be, used in this laboratory. The research context involves applied behavioral studies in which animals are trained to perform complex operant behavioral sequences, often in their homecage environment. In such studies, animals have freedom to choose whether to engage in appetitively reinforced behavioral tests that employ neither food deprivation nor fluid management. This background of operant conditioning has provided an insight to, and a context for, animal training both as an adjunct to general laboratory management and as a way to expedite scientific procedures. Thus, training has potential implications for both well-being and scientific quality, although it must be considered an adjunct to the provision of socialization with conspecifics in high quality diverse housing systems and not as an alternative to such provision. The importance of discussion and consideration of alternative procedures cannot be overemphasized.     

The technology of the Global HIV Vaccine Research Cryorepository

This paper gives an overview of our developments in the fields of cryotechnology and cryoelectronics for managing biological samples stored over liquid nitrogen, fields that are essential to a central cryostorage facility for AIDS related material used in worldwide collaborative research. As a fundamental part of this project we have developed new electronic cryovials and carrier plates attached to Flash memory chips, so that a redundant and portable set of data is present with each sample. The recovery of peripheral blood mononuclear cells (PBMCs) after cryopreservation in the new cryosubstrates showed, especially for small sample volumes (0.1 ml), an increased viability in comparison to the preservation in standard cryovials. The electronic cryosubstrates interface with a dedicated laboratory workflow management system for reproducible and standardized sample processing. This system can read standard operating procedures (SOPs) stored on each sample's memory chip. Finally, we have developed a polymorphic cryoelectronic infrastructure for sample storage tanks. This cryoelectronic system has been integrated in a hermetic storage and semiautomatic handling environment. The new technologies provide state‐of‐the‐art sample identification, documentation and tracking that brings added value to each sample. The first application of these technologies is in a worldwide collaborative research towards the production of an AIDS vaccine. The functionality and versatility of the technologies will lead to an essential optimization of sample and data exchange for global clinical studies by ensuring a high degree of standardization between laboratories.     

Genetic screening for the next decade: application of present and new technologies.

Molecular genetic technology is diffusing from the research laboratory to the clinical laboratory, where it has already begun to influence prenatal diagnosis and counseling. In the very near future, this technology will be applied more generally, using population-based screening strategies. Pilot programs are beginning to evaluate the technical feasibility and efficacy of recombinant DNA techniques for newborn screening follow-up. DNA-based population screening is being considered for heterozygous carriers of an autosomal recessive disorder such as cystic fibrosis in order to identify carrier couples at risk of having an affected child. We will review the current DNA methodologies in the context of three genetic disorders: sickle-cell disease, Duchenne muscular dystrophy, and cystic fibrosis. We will then consider the requirements for implementation of these new technologies. We will conclude that implementation will require two key factors: machines and people. Machines are required to automate molecular genetic procedures, which are currently personnel-intensive, so that the expense can be reduced and the procedures made more cost-effective. The people who are required are health professionals knowledgeable in the clinical aspects of the target disorders, as well as in the DNA laboratory testing. These professionals will be able to facilitate sample acquisition and information exchange among the laboratory, the primary health care provider, and the families requesting consultation.     

Stem cell and cellular therapy developments

Recent discoveries on human and non-human stem cells have prompted the development of several studies aimed at the translation of laboratory evidences into novel clinical procedures collectively known as ‘cellular therapy’.In this regard, a number of features specifically related to the clinical setting require stringent evaluation, including, but not limited to: ethical appropriateness; donor and recipient informed consent; autologous versus allogeneic use; media and devices for cell collection, processing, characterization, storage and distribution; donor and recipient adverse events registration and management; risk-to-benefit and cost analysis; outcome analysis; production sites accreditation and management; regulatory oversight.This article describes recent national and international developments related to the distribution of cells and tissues for clinical use. Moreover, an example is reported of the implementation of a cellular therapy production site compliant with good manufacturing practices (GMPs) in a large European University hospital.     

Inaccuracy of counts of organisms in water or other samples: effects of pre-dilution

Samples, such as raw waters, which contain large numbers of organisms will need to be pre-diluted before culture, in order to count or estimate the numbers present. This introduces a further approximation into the results obtained from routine sampling and laboratory procedures. Computers allow calculation of 95% confidence intervals (c.i.) for the estimated count in the pre-dilution sample. Although such confidence intervals can be wide, the variation in the density of organisms in the water source will be as large and probably much larger than the confidence intervals suggest. Accordingly, it is not surprisng that series of results from routine samples are highly variable.     

Sample preparation for mass spectrometry imaging: small mistakes can lead to big consequences

Mass spectrometry imaging (MSI) enables the direct analysis of molecules from the surface of a wide variety of samples, allowing the multiplex measurement of both abundance and distribution of small molecules, lipids, peptides and proteins. As the technology has been refined an increasing number of ionization methods and mass analyzers has been used that enable increased spatial and spectral resolution measurements to be made at an increased speed. Alongside the instrumentation improvements there has been optimization of sample preparation procedures that allow the highest quality data to be obtained, reproducibly, from an ever increasing diversity of samples. This review will consider the development and standardization of sample preparation methods applicable to MSI, describing the stages and procedures undertaken from the instance of sample collection, through storage, preparation and on through final processing prior to analysis. Recent technical advancements will be highlighted and areas where further experimentation and optimization may well be required will be described. All aspects of the sample preparation pipeline will be considered in detail, with examples from the literature used to emphasize why rigorous sample preparation for MSI is vital to achieve the most accurate, reproducible and validated MSI data possible.     

Use of positive reinforcement training in the management of species for reproduction

Positive reinforcement training holds great potential for enhancing the management of species for reproduction. This paper reviews a wide range of animal training activities and resultant benefits which could have application to captive reproduction programs. Recognized benefits fall into several categories. Basic animal care can be improved through voluntary cooperation by the animals with veterinary procedures and routine husbandry activities. The quality and quantity of physiological data collected for research can be improved through voluntary cooperation by the animal subjects with sample collection. Positive social interaction and reproductive behavior can be increased and aggressive behavior reduced through employment of a specialized training regime called cooperative feeding. Good success has been reported with special training programs to facilitate introduction of new members into primate social groups, while mixed results have been obtained in efforts to use training to enhance maternal skills. Finally, a discussion of both direct and indirect enhancement of psychological well-being through the use of positive reinforcement training is presented. While positive reinforcement training techniques will not be useful in every situation, information developed to date strongly suggests that these techniques should be given serious consideration when developing comprehensive programs for the management of species for reproduction. © 1994 Wiley-Liss, Inc.     

Concepts in the validation of neurochemical methods: the proper generation and use of statistics

Nearly every advance made in the study of neurotransmission has resulted from the development of new analytical methods or from the application of such methods to neurochemical problems in new ways. Each investigator places extremely high dependence on the laboratory method from which the data are gathered. It is therefore vital that these methods be proven valid when first selected and when used throughout the experiments being conducted. The process of method validation has flourished and been refined in the field of laboratory medicine. Methods are primarily validated by their accuracy and reproducibility in determining the analyte of interest in the tissue(s) to be studied. It is important that standards of performance be established that will allow objective decisions to be made when methods are tested for these characteristics. Performance standards for several neurotransmitters are suggested. Studies performed to collect method performance data are presented. From these data, statistics can be generated that help to estimate analytical errors. Guidelines for the proper generation and use of these statistics are discussed. Use of these validation approaches should be expanded in the whole of neurochemistry, which should enrich the data gathered within a laboratory and improve the harmonization between laboratories.     

Training nonhuman primates to perform behaviors useful in biomedical research

Data collected from NHPs that are trained to participate voluntarily in husbandry, veterinary, and research procedures are likely to have particular value. The authors present the results of a series of studies that examined the effects of PRT on the performance by chimpanzees of a variety of biomedically relevant behaviors: presenting their perineum for pinworm testing, providing a semen sample, presenting for an s.c. injection, and presenting for an i.m. injection. The overall trends across studies indicate that PRT techniques have significant value in the handling and management of NHPs in many laboratory research settings, including less variability in the data collected and fewer potential confounding variables, which should lead to important refinements in the definition of NHPs as biomedical research models.     

In vitro evaluation of sperm quality

This paper highlights selected laboratory analyses that are currently used to evaluate sperm, and describes why results from these assays do not consistently correlate with sperm fertility. Reasons for the disconnect between the two are due in part to the definition and reliability of the fertility data collected, to the complexity of the spermatozoon itself, to imprecision of some measurements, and to uncontrollable factors not associated to either the laboratory analysis or the sperm sample. Each sperm must possess a number of different attributes to fertilize an oocyte, and individual laboratory assays measure only one or a few of these attributes. Current and past data, correlating laboratory assay data with sperm fertility are presented in an effort to determine which types of assays are important to conduct and when to conduct them. Even though laboratory assay results do not allow accurate evaluation of the fertilizing potential of a semen sample, these assays are important to enable culling of poor quality samples.     

Harmonisation of Measurement Procedures: how do we get it done?

Clinical laboratory measurement results must be comparable among different measurement procedures, different locations and different times in order to be used appropriately for identifying and managing disease conditions. Harmonisation in the broad sense is the overall process of achieving comparability of results among clinical laboratory measurement procedures that measure the same measurand. The term standardisation is used when comparable results among measurement procedures are based on calibration traceability to SI using a reference measurement procedure of the highest available order. When there is no higher order reference measurement procedure available, and it is unlikely that one can be developed, the term harmonisation refers to any process for achieving comparable results among measurement procedures for an individual measurand.This review explains calibration traceability and focuses on the principles of harmonisation for those measurands for which a reference measurement procedure does not exist. We discuss the value of harmonisation, the importance of commutable reference materials, the barriers to harmonisation that exist today, and conclude with a discussion of a current global effort to improve the state of harmonisation.