Use of Modified Fraser's Stain in Promoting Activity Test (PAT)

The Promoting Activity Test (PAT) requires a staining procedure that allows rapid, accurate and reliable counting of mitotic figures. We propose use of Fraser's kernechtrotcrystal violet technique, but eliminating the picric-alcoholic differentiation to avoid fading. This modified protocol gives higher mitotic counts in adult mouse adrenal cortex than die hematoxylin-eosin originally used, especially with respect to less conspicuous prophases.     

Interactive Multimedia for Promoting Physical Activity (IMPACT) in Children

Objective: To develop and examine the efficacy of a computer‐based interactive multimedia curriculum for promoting physical activity in fourth grade children. Research Methods and Procedures: The participants were 209 fourth grade children (mean age of 9.5 ± 0.4 years) from four schools. Two schools received an 8‐week multimedia intervention delivered by interactive CD‐ROM, supplemented by four classroom and four homework assignments. Two control schools received educational CD‐ROMS not related to health outcomes. Measures conducted before and after intervention included height, weight, percentage body fat (bioimpedance analysis), physical activity (5‐day accelerometry), and psychosocial aspects of physical activity by questionnaire. All outcomes were examined using general linear models. Results: There was a significant treatment effect for obesity reduction in girls but not in boys. There were no significant treatment effects on total physical activity by accelerometry (total counts per minute), but there was an overall treatment effect on reducing percent of time in moderate‐intensity activity (16.5% to 15% of the time) and significant sex‐by‐ treatment interactions for light‐intensity activities (reduction in boys from 78% to 75% of the time and an increase in girls from 78% to 81% of the time). There were marginal/significant treatment effects for improvements in behavioral outcomes, including self‐efficacy (p = 0.06), social norms (p = 0.07), and outcome expectancies (p = 0.049). Discussion: The interactive multimedia curriculum favored an improvement in obesity indices in girls and was associated with subtle changes in physical activity in girls and general improvement in psychosocial outcomes related to physical activity.     

Use of the Gram stain in microbiology

The Gram stain differentiates bacteria into two fundamental varieties of cells. Bacteria that retain the initial crystal violet stain (purple) are said to be 'Gram-positive,' whereas those that are decolorized and stain red with carbol fuchsin (or safranin) are said to be 'Gram-negative.' This staining response is based on the chemical and structural makeup of the cell walls of both varieties of bacteria. Gram-positives have a thick, relatively impermeable wall that resists decolorization and is composed of peptidoglycan and secondary polymers. Gram-negatives have a thin peptidoglycan layer plus an overlying lipid-protein bilayer known as the outer membrane, which can be disrupted by decolorization. Some bacteria have walls of intermediate structure and, although they are officially classified as Gram-positives because of their linage, they stain in a variable manner. One prokaryote domain, the Archaea, have such variability of wall structure that the Gram stain is not a useful differentiating tool.     

Use of the Gram stain in microbiology

The Gram stain differentiates bacteria into two fundamental varieties of cells. Bacteria that retain the initial crystal violet stain (purple) are said to be ''Gram-positive,'' whereas those that are decolorized and stain red with carbol fuchsin (or safranin) are said to be ''Gram-negative.'' This staining response is based on the chemical and structural makeup of the cell walls of both varieties of bacteria. Gram-positives have a thick, relatively impermeable wall that resists decolorization and is composed of peptidoglycan and secondary polymers. Gram-negatives have a thin peptidoglycan layer plus an overlying lipid-protein bilayer known as the outer membrane, which can be disrupted by decolorization. Some bacteria have walls of intermediate structure and, although they are officially classified as Gram-positives because of their linage, they stain in a variable manner. One prokaryote domain, the Archaea, have such variability of wall structure that the Gram stain is not a useful differentiating tool.     

Complete genome sequence of Ferrimonas balearica type strain (PAT)

Ferrimonas balearica Rossello-Mora et al. 1996 is the type species of the genus Ferrimonas, which belongs to the family Ferrimonadaceae within the Gammaproteobacteria. The species is a Gram-negative, motile, facultatively anaerobic, non spore-forming bacterium, which is of special interest because it is a chemoorganotroph and has a strictly respiratory metabolism with oxygen, nitrate, Fe(III)-oxyhydroxide, Fe(III)-citrate, MnO(2), selenate, selenite and thiosulfate as electron acceptors. This is the first completed genome sequence of a member of the genus Ferrimonas and also the first sequence from a member of the family Ferrimonadaceae. The 4,279,159 bp long genome with its 3,803 protein-coding and 144 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.     

A preliminary study of the wild siamang gibbon (Hylobates syndactylus) at Fraser's Hill,Malaysia

The wild siamang gibbon was studied at Fraser's Hill, Malaysia. The study area was covered with a well developed forest and offered a suitable habitat for siamang gibbons. Other primates living in this area were the white-handed gibbon, the duskey leafmonkey, the banded leaf-monkey, and the pig-tailed monkey. The siamang gibbon groups observed have a monogamous structure consisting of one pair of adult individuals and one or more subadults which are assumed to be the offspring of the adults. The adult male showed behavior typical of a group leader. As subadults become older, they tend to become spatially separated from the mother group. Each group was observed to range freely within an exclusive area of the forest into which no other group was observed to intrude. Each group emitted loud vocalizations which seemed to maintain the spacing between the groups.     

Process analytical technology (PAT) tools for the cultivation step in biopharmaceutical production

The process analytical technology (PAT) initiative is now 10 years old. This has resulted in the development of many tools and software packages dedicated to PAT application on pharmaceutical processes. However, most applications are restricted to small molecule drugs, mainly for the relatively simple process steps like drying or tableting where only a limited number of parameters need to be controlled. A big challenge for PAT still lies in applications for biopharmaceuticals and then especially in the cultivation process step, where the quality of a biopharmaceutical product is largely determined. This review gives an overview of the currently available tools for monitoring and controlling the biopharmaceutical cultivation step and of the main challenges for the most common cell platforms (i.e. Escherichia coli, yeast, and mammalian cells) used in biopharmaceutical manufacturing. The real challenge is to understand how intracellular mechanisms (from synthesis to excretion) influence the quality of biopharmaceuticals and how these mechanisms can be monitored and controlled to yield the desired end product quality. Modern “omics” tools and advanced process analyzers have opened up the way for PAT applications for the biopharmaceutical cultivation process step.     

Assessment of Phosphinothricin Acetyltransferase (PAT) Degradation From Transgenic Zoysiagrass Digested with Simulated Gastric Fluid (SGF)

The in vitro pepsin digestion assay is the international standard for assessing the safety or risk of novel proteins newly produced in transgenic crops. However, this protocol, based on the degradation of protein purified from Escherichia coli, has recently been criticized for problems such as its objective detection limit. Here, we estimated the digestion stability of the phosphinothricin acetyltransferase (PAT) protein in soluble proteins as well as from leaf tissue powder in simulated gastric fluid (SGF). Our line of genetically modified zoysiagrass carried a single copy of the bar gene, which entered a chromosomal region not encoding protein. We designated it as Jeju Green 21 (JG21). From total soluble proteins extracted from JG21 leaves, digestibility of the PAT protein in SGF was examined by enzymatic assays, sodium dodecyl sulfate–polyacrylamide gel electrophoresis, Western gel blots, and an immunodetection strip kit. The degradability of pure PAT protein obtained from E. coli was clearly apparent within at least 30 s. However, PAT degradation in leaf tissue powder was significantly delayed, indicating that some matrices in that powder might have influenced its digestion stability by SGF. Nevertheless, degradation of the powder (real-life) sample was complete within at least 5 min, suggesting that this protein produced in JG21 zoysiagrass can be digested harmlessly in the stomachs of humans or livestock.     

Use of an improved zirconyl hematoxylin stain in the diagnosis of Barrett's esophagus

Barrett's esophagus is a precancerous condition characterized by replacement of the normal stratified squamous epithelium by a simple columnar epithelium with goblet cells that secrete an acidic mucin. As originally formulated, fresh solutions of zirconyl hematoxylin stain goblet cells poorly. An improved formula, quintupling the amount of oxidant, yields zirconyl hematoxylin solutions that stain goblet cells darkly even when fresh. The improved zirconyl hematoxylin can be used in place of alcian blue in the diagnosis of Barrett's esophagus. The ingredients of zirconyl hematoxylin are always readily available and are generally recognized as safe.     

Case study and application of process analytical technology (PAT) towards bioprocessing: II. Use of ultra-performance liquid chromatography (UPLC) for making real-time pooling decisions for process chromatography

Process analytical technology (PAT) has been gaining a lot of momentum in the biopharmaceutical community due to the potential for continuous real-time quality assurance resulting in improved operational control and compliance. Two of the key goals that have been outlined for PAT are "variability is managed by the process" and "product quality attributes can be accurately and reliably predicted over the design space established for materials used, process parameters, manufacturing, environmental, and other conditions". Recently, we have been examining the feasibility of applying different analytical tools for designing PAT applications for bioprocessing. We have previously shown that a commercially available online high performance liquid chromatography (HPLC) system can be used for analysis that can facilitate real-time decisions for column pooling based on product quality attributes (Rathore et al., 2008). In this article we test the feasibility of using a commercially available ultra- performance liquid chromatography (UPLC) system for real-time pooling of process chromatography columns. It is demonstrated that the UPLC system offers a feasible approach and meets the requirements of a PAT application. While the application presented here is of a reversed phase assay, the approach and the hardware can be easily applied to other modes of liquid chromatography.     

Cis-dichloro-diammine platinum (II) as stain for electron microscopic preparations

Summary Cis-dichloro-diammine platinum (II) provides a strong contrast to electron microscopic preparations by reacting with nucleic acids and proteins. This staining technique is easy and applicable to ultrathin sections embedded in Epon; various staining conditions have been tested.