Standardized staining methods: Feulgen-Rossenbeck reaction for desoxyribonucleic acid and periodic acid-Schiff (PAS) procedure

A project group working under the European Confederation of Laboratory Medicine (ECLM) presents recommendations for standardized procedures for the Feulgen-Rossenbeck-Schiff and the periodic acid-Schiff (PAS) reactions on cytological and histological material. The advantages and disadvantages of such standardized procedures are presented here in a preamble. Both users and manufacturers are encouraged to give their opinions with a view to achieving consensus on these procedures and on how further work on these lines may proceed.     

Standardization of procedures for nitrogen fractionation of ruminant feeds

The Cornell Net Carbohydrate Protein Model (Chalupa et al., 1991; Sniffen et al., 1992) has developed the need for uniform procedures to partition feed nitrogen into A, B, and C fractions (Pichard and Van Soest, 1977). While carbohydrate fractions are relatively standardized (based on NDF, ADF with corrections for ash, protein, and lignin), the fractionation of plant nitrogen has been open to considerable variation in procedures. This has led to non-uniformity among reported values for nitrogen fractions. This paper recommends reliable procedures for nonprotein nitrogen (NPN) and buffer-soluble protein. These procedures have been examined for reproducibility and relevance to biological expectations. Procedures for acid-detergent insoluble nitrogen (ADIN), and neutral-detergent insoluble nitrogen (NDIN) are also included as they are required for the model. Some alternatives in certain procedures are offered.     

Further studies on the cytochemistry of the standardized silver staining of interphase nucleoli in smear preparations of Yoshida ascitic sarcoma cells in rats

Silver staining procedure for the selective demonstration of nucleolar silver stained granules (SSG) and for the simultaneous demonstration of SSG and nucleolar silver stained matrix (SSM) were studied in smears of rat Yoshida sarcoma cells. The successful results of these procedures depend mainly on the quality of silver nitrate and formaldehyde. However, both chemicals can be easily standardized and stabilized disregarding their origin and batch. In standardized procedures (one-step procedure for the selective demonstration of SSG and two-steps procedure for the simultaneous demonstration of SSG and SSM) the silver is apparently bound to acidic groups of proteins of SSG and SSM. The proteins of SSG and SSM seem to be different but both belong to the group of acidic non-histone proteins. According to the results of digestion experiments a possibility also exists that the acidic proteins of SSG may be associated with DNA. The identification of SSG visualized by described standardized procedures was determined not only by cytochemical extraction tests but also by biological experiments. The latter demonstrated that the number of SSG in Yoshida sarcoma cells decreases after treatment of experimental animals with actinomycin D and therefore depends on the state of the nucleolar RNA synthesis.     

Microbiological evaluation of a biological safety cabinet modified for bedding disposal

A biological safety cabinet modified for bedding disposal was tested to determine the cabinet's ability to protect operators and experiments from aerosol exposure during routine microbiological and cage cleaning procedures. Stringent test conditions were provided by modifications of standardized protocols in addition to simulated cage dumping procedures, both of which utilized bacterial aerosols as challenges. Results of standardized test procedures (with no operator present) indicated good performance in protecting both operators and experiments. Procedures involving the dumping (by an operator) of contaminated bedding within the unit showed that the cabinet was able to contain 99.96% or greater of the total particles generated.     

Further studies on the cytochemistry of the standardized silver staining of interphase nucleoli in smear preparations of yoshida ascitic sarcoma cells in rats

Summary Silver staining procedure for the selective demonstration of nucleolar silver stained granules (SSG) and for the simultaneous demonstration of SSG and nucleolar silver stained matrix (SSM) were studied in smears of rat Yoshida sarcoma cells. The successful results of these procedures depend mainly on the quality of silver nitrate and formaldehyde. However, both chemicals can be easily standarized and stabilized disregarding their origin and batch. In standardized procedures (one-step procedure for the selective demonstration of SSG and two-steps procedure for the simultaneous demonstration of SSG and SSM) the silver is apparently bound to acidic groups of proteins of SSG and SSM. The proteins of SSG and SSM seem to be different but both belong to the group of acidic non-histone proteins. According to the results of digestion experiments a possibility also exists that the acidic proteins of SSG may be associated with DNA. The identification of SSG visualized by described standardized procedures was determined not only by cytochemical extraction tests but also by biological experiments. The latter demonstrated that the number of SSG in Yoshida sarcoma cells decreases after treatment of experimental animals with actinomycin D and therefore depends on the state of the nucleolar RNA synthesis.     

Standardized Viral Hemagglutination and Hemagglutination-Inhibition Tests. II. Description and Statistical Evaluation

Standardized hemagglutination and hemagglutination-inhibition procedures are described and statistically evaluated for all animal viruses where applicable, except for rubella and the arbovirus group. The standardized tests employ a constant phosphate-buffered saline diluent and constant volumes of serum, antigen, and standardized erythrocyte suspension. The standardized hemagglutination test has a reproducibility of 84 to 96% with adenoviruses, rubeola, and the myxoviruses, and 78 to 93% with reoviruses; the standardized hemagglutination-inhibition test has a reproducibility of 95 to 100% with all viruses tested.     

Protocol for X-ray dosimetry and exposure arrangements employed in studies of late somatic effects in mammals

A number of European laboratories studying the late effects of ionizing radiation in animals have established an effective cooperation within the European Late Effects Project Group (EULEP) since 1970. To facilitate the exchange of biological results several techniques, including quality control of the experimental animals, pathology and dosimetry, have to be standardized. The most important aspects of the procedures for X-irradiation and dosimetry of small animals are summarized. These include recommendations on irradiation conditions, dosimetry methods, characteristics of phantoms and factors affecting X-ray dosimetry. X-irradiation procedures employed by the participating institutes are described and the results of five X-ray dosimetry intercomparisons are reported. The introduction of a common dosimetry protocol has resulted in improvements in exposure arrangements and absolute dosimetry.     

The physical attractiveness of facially deformed patients before and after craniofacial surgery

The present experiment investigated whether the physical attractiveness of craniofacially deformed children and adolescents could be improved by surgical procedures. Twenty patients between the ages of 5 months and 17 years were randomly selected from patient files. Patient diagnoses included facial clefts, hypertelorism, Treacher Collins syndrome, and craniofacial dysostosis (Crouzon's and Apert's syndromes). Rigorously standardized photographs of patients taken before and after surgery were shown to 40 "naive" raters ranging in age from 17 to 52 years. Raters analyzed the photographs with regard to global physical attractiveness. These ratings indicated that the patients' physical attractiveness was reliably (62 percent) improved following surgery. The results are discussed in light of recent evidence that untreated craniofacial patients may be at risk for psychosocial disorders and in terms of the growing evidence of the importance of physical appearance for the development of cognitive and social-emotional competence. In addition, a standardized assessment system is described that can be used to facilitate the compilation of actuarial data predicting surgical outcomes. Finally, the importance of empirically evaluating the effectiveness of surgical procedures and practitioners on a continuing basis is emphasized.     

Applications of MALDI-TOF mass spectrometry in clinical diagnostic microbiology

Until recently, microbial identification in clinical diagnostic laboratories has mainly relied on conventional phenotypic and gene sequencing identification techniques. The development of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) devices has revolutionized the routine identification of microorganisms in clinical microbiology laboratories by introducing an easy, rapid, high throughput, low-cost, and efficient identification technique. This technology has been adapted to the constraint of clinical diagnostic laboratories and has the potential to replace and/or complement conventional identification techniques for both bacterial and fungal strains. Using standardized procedures, the resolution of MALDI-TOF MS allows accurate identification at the species level of most Gram-positive and Gram-negative bacterial strains with the exception of a few difficult strains that require more attention and further development of the method. Similarly, the routine identification by MALDI-TOF MS of yeast isolates is reliable and much quicker than conventional techniques. Recent studies have shown that MALDI-TOF MS has also the potential to accurately identify filamentous fungi and dermatophytes, providing that specific standardized procedures are established for these microorganisms. Moreover, MALDI-TOF MS has been used successfully for microbial typing and identification at the subspecies level, demonstrating that this technology is a potential efficient tool for epidemiological studies and for taxonomical classification.     

Collecting Saliva and Measuring Salivary Cortisol and Alpha-amylase in Frail Community Residing Older Adults via Family Caregivers

Salivary measures have emerged in bio-behavioral research that are easy-to-collect, minimally invasive, and relatively inexpensive biologic markers of stress. This article we present the steps for collection and analysis of two salivary assays in research with frail, community residing older adults-salivary cortisol and salivary alpha amylase. The field of salivary bioscience is rapidly advancing and the purpose of this presentation is to provide an update on the developments for investigators interested in integrating these measures into research on aging. Strategies are presented for instructing family caregivers in collecting saliva in the home, and for conducting laboratory analyses of salivary analytes that have demonstrated feasibility, high compliance, and yield quality specimens. The protocol for sample collection includes: (1) consistent use of collection materials; (2) standardized methods that promote adherence and minimize subject burden; and (3) procedures for controlling certain confounding agents. We also provide strategies for laboratory analyses include: (1) saliva handling and processing; (2) salivary cortisol and salivary alpha amylase assay procedures; and (3) analytic considerations.     

New method for evaluation of virucidal activity of antiseptics and disinfectants.

Counting culturable viruses adsorbed to cellulose nitrate filters (the VIRADEN method) is proposed as a simple procedure for the evaluation of the virucidal activity of antiseptics and disinfectants. The virucidal activities of two different doses of iodine, chlorine, glutaraldehyde, and chlorhexidine digluconate on poliovirus 1 were tested with a standardized procedure and with the VIRADEN method. The two procedures assayed provided similar results.     

New Method for Evaluation of Virucidal Activity of Antiseptics and Disinfectants

Counting culturable viruses adsorbed to cellulose nitrate filters (the VIRADEN method) is proposed as a simple procedure for the evaluation of the virucidal activity of antiseptics and disinfectants. The virucidal activities of two different doses of iodine, chlorine, glutaraldehyde, and chlorhexidine digluconate on poliovirus 1 were tested with a standardized procedure and with the VIRADEN method. The two procedures assayed provided similar results.     

Bridging molecular genetics and participatory research: how access and benefit‐sharing stimulate interdisciplinary research for tropical biology and conservation

Molecular genetics research can benefit efforts to conserve the genetic diversity of tropical plant species. Clear and efficient procedures are needed to access DNA samples, while respecting tropical countries’ and local communities’ rights on genetic resource usage. The Nagoya Protocol on Access and Benefit‐Sharing, which took effect in 2014, provides an opportunity to establish such procedures. However, scientists are concerned that its emphasis on monetary gains restricts research focused on scientific, societal, and environmental benefits. Despite much political and scientific debate, few concrete cases have demonstrated the practical functioning of the Nagoya Protocol. This paper describes the first application of the Protocol in Guatemala, where it was used to grant permission to a non‐commercial study on gene flow in mahogany (Swietenia macrophylla King) populations in the Maya Biosphere Reserve of Petén. On the basis of this study, we discuss five strategies to enhance the application of molecular genetics to conservation biology under the Nagoya Protocol: (1) generate short and standardized procedures; (2) enable science communication; (3) cultivate a common understanding between users, providers, and potential beneficiaries; (4) involve local research and practitioner organizations; and (5) integrate participatory research. Positive societal views on the application of molecular genetics to conservation biology generate further support for work in this discipline and promote adoption of research results for the conservation of genetic diversity of tropical plant species.     

Tests de Selection des Spermatozoïdes avant Assistance Médicale à la Procréation

There are many procedures to select spermatozoa but those procedures are often specific of each andrology centre. This review that is not an exhaustive review of the numerous methods published is rather devoided to focus on the main techniques, the sperm washing, the filtration methods («SpermPrep»), the swim-up, and the discontinuous density gradients with colloïdal solutions («Percoll», PureSperm” and «Isolate») including iodinated organic molecule as ‘OptiPrep». The first part presents a synthesis of literature in order to bring out standardized procedures, and a summary of the advantages and disadvantages of each step of these procedures. Some specific anomalies of sperm need particular treatments. There is a consensus on the validated techniques to prepare semen samples in the presence of antisperm antibodies, leukocytes, bacterial contaminants, and also semen samples exhibiting hyperviscosity or in case of retrograde ejaculation. Cryopreserved sperm preparation is also described with references to experimental results in freezing-thawing human spermatozoa. The second part proposes guidelines for the therapeutic choice of the appropriate assisted reproductive technologies (i.e.: intra uterine artificial insemination, in vitro fertilization or intracytoplasmic sperm injection) according to semen parameters, and to the outcome of sperm selection and survival.     

Comparison of Methods for Coccidioidomycosis Complement Fixation

A Laboratory Branch Task Force of the National Communicable Disease Center has proposed a standardized complement fixation procedure (LBCF) and an adaptation of this to microtitration techniques (MT) as uniform methods for performing complement fixation (CF) tests. A common procedure should make CF results from one laboratory more comparable to another. In addition, it would be preferable if the common procedure reproduced the titer levels of a testing procedure which is to be replaced, particularly when valid clinical interpretations have been derived from the latter. Replicated sets of sera were tested by the LBCF, MT, and the standard Smith CF procedure for coccidioidomycosis. Results with all three procedures were highly reproducible within an acceptable one-tube variation of a twofold dilution series, but the frequency of one-tube variations was greater with the MT method than with the other two. There was no statistical difference in the titers obtained with the Smith and LBCF procedures, but there was a significant difference when the MT results were compared to those with the Smith method. The LBCF method should be acceptable as a standardized and uniform CF procedure for coccidioidomycosis, subject to comparative testing between different laboratories.     

Pre-analytical operating procedures for serum Low Molecular Weight protein profiling

Biological specimen collection and storage are an integral component of serum proteomics research. Although many efforts have been posed to address the effects of pre-analytical procedures, standardized protocols for collection and storage of samples for Low Molecular Weight (LMW) proteome profiling are still needed.Here we report a systematic analysis on the influence of pre-analytical factors [clotting times, temperature and time storage, addition of protease inhibitor (PI)] on serum LMW proteome profiling. Moreover, a comparison between manual versus automated peptide purification by functionalized magnetic bead-based MALDI-MS approach was performed. The results demonstrated best serum LMW proteins recovery and stability using a clotting time between 1 and 2 h, with serum stored up to 2 h either at room temperature or at 4 °C, independently of PI addition. PI addition to whole blood resulted in a lower number of LMW peaks detected. Finally, minimal effects on serum proteome profiles were observed after 1-month storage at ? 80 °C, independently of PI addition on whole blood and/or serum.In conclusion, the use of standardized pre-analytical and storage procedures together with an automated peptide purification might minimize potential bias on serum LMW profiling results, thus allowing a better homogeneity and reproducibility in future proteomics studies.     

Biomarker evaluation and comparison using the controls as a reference population

The classification accuracy of a continuous marker is typically evaluated with the receiver operating characteristic (ROC) curve. In this paper, we study an alternative conceptual framework, the "percentile value." In this framework, the controls only provide a reference distribution to standardize the marker. The analysis proceeds by analyzing the standardized marker in cases. The approach is shown to be equivalent to ROC analysis. Advantages are that it provides a framework familiar to a broad spectrum of biostatisticians and it opens up avenues for new statistical techniques in biomarker evaluation. We develop several new procedures based on this framework for comparing biomarkers and biomarker performance in different populations. We develop methods that adjust such comparisons for covariates. The methods are illustrated on data from 2 cancer biomarker studies.     

Construction of a Preclinical Multimodality Phantom Using Tissue-mimicking Materials for Quality Assurance in Tumor Size Measurement

World Health Organization (WHO) and the Response Evaluation Criteria in Solid Tumors (RECIST) working groups advocated standardized criteria for radiologic assessment of solid tumors in response to anti-tumor drug therapy in the 1980s and 1990s, respectively. WHO criteria measure solid tumors in two-dimensions, whereas RECIST measurements use only one-dimension which is considered to be more reproducible ^{1, 2, 3,4,5}. These criteria have been widely used as the only imaging biomarker approved by the United States Food and Drug Administration (FDA) ⁶. In order to measure tumor response to anti-tumor drugs on images with accuracy, therefore, a robust quality assurance (QA) procedures and corresponding QA phantom are needed.To address this need, the authors constructed a preclinical multimodality (for ultrasound (US), computed tomography (CT) and magnetic resonance imaging (MRI)) phantom using tissue-mimicking (TM) materials based on the limited number of target lesions required by RECIST by revising a Gammex US commercial phantom ⁷. The Appendix in Lee et al. demonstrates the procedures of phantom fabrication ⁷. In this article, all protocols are introduced in a step-by-step fashion beginning with procedures for preparing the silicone molds for casting tumor-simulating test objects in the phantom, followed by preparation of TM materials for multimodality imaging, and finally construction of the preclinical multimodality QA phantom. The primary purpose of this paper is to provide the protocols to allow anyone interested in independently constructing a phantom for their own projects. QA procedures for tumor size measurement, and RECIST, WHO and volume measurement results of test objects made at multiple institutions using this QA phantom are shown in detail in Lee et al. ⁸.     

Chemical Approaches for Stabilizing Perovskite Solar Cells

Chemical bonding dictates not only the optoelectronic properties of materials, but also the intrinsic and extrinsic stability of materials. Here, the causes of intrinsic and extrinsic instability of perovskite materials are reviewed considering their correlation with the unique chemical‐bonding nature of perovskite materials. There are a number of key standardized stability tests established by the International Electrotechnical Commission for commercialized photovoltaic modules. Based on these procedures, the possible causes and related mechanisms of the material degradation that can arise during the test procedures are identified, which are discussed in terms of their chemical bonds. Based on the understanding of the critical causes, promising strategies for mitigating the causes to enhance the stability of perovskite solar cells are summarized. The stability of the state‐of‐the‐art perovskite solar cells implies a need for the development of improved stability‐testing protocols to move onto the next stage toward commercialization.     

Techniques for physicochemical characterization of nanomaterials

Advances in nanotechnology have opened up a new era of diagnosis, prevention and treatment of diseases and traumatic injuries. Nanomaterials, including those with potential for clinical applications, possess novel physicochemical properties that have an impact on their physiological interactions, from the molecular level to the systemic level. There is a lack of standardized methodologies or regulatory protocols for detection or characterization of nanomaterials. This review summarizes the techniques that are commonly used to study the size, shape, surface properties, composition, purity and stability of nanomaterials, along with their advantages and disadvantages. At present there are no FDA guidelines that have been developed specifically for nanomaterial based formulations for diagnostic or therapeutic use. There is an urgent need for standardized protocols and procedures for the characterization of nanoparticles, especially those that are intended for use as theranostics.