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Working relationships between vendors and academic libraries have extended from a traditional one of customer and bookseller to the testing of vendor-developed software. This extension has occurred since 1980, the last quarter century. What has changed since 2000 is the addition of tests of vendor/client interface informed by enterprise software that completely alters methodologies for accommodating library requirements for book supply, MARC record supply, and electronic invoices and purchase orders. This paper will examine the technical, administrative, and legal considerations now relevant to beta testing of vendor software by library staff and offer charts and checklists that may make these projects more easily comprehended. [Copyright &y& Elsevier]

A number of enzyme-linked immunosorbent assays (ELISAs) have been developed for the detection of milk adulteration in dairy products. Target antigens have been caseins, lactoglobulins, immunoglobulins and other whey proteins. Polyclonal and monoclonal antibodies have been used in a variety of formats including direct, indirect, competitive and sandwich ELISAs. ELISAs have been successfully applied to the detection of cows' milk adulteration of sheep, goat and buffalo milk. Goat milk adulteration of sheep milk has also been detected. A number of ELISAs have also been applied to cheese. It is recommended that ELISA should be used in combination with PCR to ensure compliance with current legislation. [ABSTRACT FROM AUTHOR]

This paper describes the development and use of a competitive enzyme-linked immunosorbent assay (ELISA) to detect prawns which have been irradiated. The ELISA utilizes a monoclonal antibody against a modified DNA base, dihydrothymidine. A comparison of extraction procedures demonstrated that DNA purification was not required and that crude prawn homogenate could be used in the ELISA. The ELISA was applied successfully to two prawn species, North Atlantic prawn (Pandalus borealis) and Tiger prawn (Penaeus monodon). The ELISA has a working range of 0.5–2 kGy with CVs typically below 10%. Storage of irradiated prawns for up to 12 months at -20 °C had no effect on ELISA performance. As most food contains DNA the assay has potential to be applied in a wide range of foodstuffs. [ABSTRACT FROM AUTHOR]

Heat shock proteins (Hsps) are known to be induced in response to short-term stress. The present study aimed to evaluate the potential of Hsp70 as a biomarker of stress produced by increased temperature, osmotic pressure, and exposure to cadmium and sodium chloride in marine macroalgae and fresh water plant species. An indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) was developed with a working range of 0.025-10 μg ml -1 using a monoclonal antibody raised against purified Hsp70 of Phaseolus aureus (mung bean). Fucus serratus (toothed wrack), Chondrus crispus (Stackhouse or Carrageen moss), Ulva lactuca (sea lettuce) and Lemna minor (common duckweed) sample extracts were stressed for up to 24 h and then tested in the IC-ELISA. The presence of Hsp70 and cross-reactivity of the monoclonal antibody was confirmed by Western blot. The heat shock response was confirmed in each species using a 2-h 42°C treatment. Following heat shock, Hsp70 concentrations increased to a peak at 2 h ( F. serratus ) or 4 h ( L. minor ), after which concentrations decreased. Osmotic and cadmium stresses also resulted in elevated Hsp70 concentrations in samples of F. serratus and L. minor when compared with unstressed controls. In both, osmotic and metal stress, the production of Hsp70 increased to a maximum and subsequently decreased as the stressor levels increased. Results suggest that Hsp70 IC-ELISA could potentially be applied to the detection of stress in these aquatic species, although it would probably be most effective when used in conjunction with other measurements to provide a stressor-specific biomarker profile or fingerprint. [ABSTRACT FROM AUTHOR]

Background: Patients with implantable cardioverter–defibrillators often receive adjunctive antiarrhythmic therapy to prevent frequent shocks. We tested the efficacy and safety of sotalol, a beta-blocker with class III antiarrhythmic effects, for this purpose. Methods: In a multicenter trial, patients were stratified according to left ventricular ejection fraction (≤0.30 or >0.30), randomly assigned to double-blind treatment with 160 to 320 mg of sotalol per day (151 patients) or matching placebo (151 patients), and followed for 12 months. Kaplan–Meier analyses of the time to an event were performed. Three end points were used: the delivery of a first shock for any reason or death from any cause, the first appropriate shock for a ventricular arrhythmia or death from any cause, and the first inappropriate shock for a supraventricular arrhythmia or death from any cause. Results: Compliance with double-blind treatment was similar in the two groups. There were seven deaths in the placebo group and four in the sotalol group. As compared with placebo, treatment with sotalol was associated with a lower risk of death from any cause or the delivery of a first shock for any reason (reduction in risk, 48 percent; P<0.001 by the log-rank test), death from any cause or the delivery of a first appropriate shock (reduction in risk, 44 percent; P=0.007), or death from any cause or the delivery of a first inappropriate shock (reduction in risk, 64 percent; P=0.004). Sotalol also reduced the mean (±SD) frequency of shocks due to any cause (1.43± 3.53 shocks per year, as compared with 3.89±10.65 in the placebo group; P=0.008). In the sotalol group, the reduction in the risk of death from any cause or the delivery of a first shock for any reason did not differ significantly between patients with ejection fractions of more than 0.30 and those with ejection fractions of 0.30 or less. Conclusions: Oral sotalol was safe and efficacious in reducing the risk of death or the delivery of a first defibrillator shock whether or not ventricular function was depressed. (N Engl J Med 1999;340:1855-62.) [ABSTRACT FROM AUTHOR]

The article presents a kinetic study on the reaction between superoxide and cobalamins II (Cbl (II)) and on the ability of cyanocobalamin, the vitamin B12 that forms in vitamin pills, in preventing intracellular superoxide levels in a cell. It notes that all of the kinetic experiments were made in the presence of catalase. It cites that the study has demonstrated that the intracellular Cbl form cob(II)alamin reacts with superoxide at rates nearing those of superoxide dismutase (SOD).

Endogenous molecules, such as heat shock proteins (HSP), can function as danger signals when released into the extracellular environment in response to cell stress, where they elicit an immune response such as cytokine secretion. There has also been some suggestion that contamination of exogenous HSPs with lipopolysaccharide (LPS) may be responsible for these effects. This study investigates the effects of exogenous HSPA1A and HSPB1 on the activation of immune cells and the resulting secretion of cytokines, which are involved in inflammatory responses. To address whether exogenous HSPs can directly activate cytokine secretion, naïve U937 cells, differentiated U937 cells and peripheral blood mononuclear cells (PBMCs) were treated with either exogenously applied HSPA1A or HSPB1 and then secreted IL-1β, TNF-α and IL-10 were measured by ELISA. Both HSPs were able to induce a dose-dependent increase in IL-10 secretion from naïve U937 cells and dose-dependent IL-1β, TNF-α and IL-10 secretion were also observed in differentiated U937 cells and PBMCs. We also observed that CD14 affects the secretion levels of IL-1β, TNF-α and IL-10 from cells in response to exogenous HSP treatment. In addition, HSPA1A and HSPB1 were shown to interact with CD14, CD36 and CD11b extracellular receptor proteins. Several approaches used in this study indicate that HSP-induced cytokine secretion is largely independent of any contaminating LPS in the samples., (© 2023 The Authors. FEBS Open Bio published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Preclinical studies are an essential stage for any pharmacological agent hoping to make its way into clinical trials. An ideal preclinical model that can accurately predict clinical response does not exist and the best that the scientific community have at the moment is to select the most relevant study model pertaining to the disease of interest from those available, which includes: cell lines, animal models, and even in-silico methodology. Currently, there is a huge gap between preclinical and clinical trial results, indicating that there is much room for improvement in developing a better model to bridge the translational gap.

HSPC1 is a critical protein in cancer development and progression, including colorectal cancer (CRC). However, clinical trial data reporting the effectiveness of HSPC1 inhibitors on several cancer types has not been as successful as predicted. Furthermore, some N-terminal inhibitors appear to be much more successful than others despite similar underlying mechanisms. This study involved the application of three N-terminal HSPC1 inhibitors, 17-DMAG, NVP-AUY922 and NVP-HSP990 on CRC cells. The effects on client protein levels over time were examined. HSPC1 inhibitors were also applied in combination with chemotherapeutic agents commonly used in CRC treatment (5-fluorouracil, oxaliplatin and irinotecan). As HSPA1A and HSPB1 have anti-apoptotic activity, gene-silencing techniques were employed to investigate the significance of these proteins in HSPC1 inhibitor and chemotherapeutic agent resistance. When comparing the action of the three HSPC1 inhibitors, there are distinct differences in the time course of important client protein degradation events. The differences between HSPC1 inhibitors were also reflected in combination treatment-17-DMAG was more effective compared with NVP-AUY922 in potentiating the cytotoxic effects of 5-fluorouracil, oxaliplatin and irinotecan. This study concludes that there are distinct differences between N-terminal HSPC1 inhibitors, despite their common mode of action. Although treatment with each of the inhibitors results in significant induction of the anti-apoptotic proteins HSPA1A and HSPB1, sensitivity to HSPC1 inhibitors is not improved by gene silencing of HSPA1A or HSPB1. HSPC1 inhibitors potentiate the cytotoxic effects of chemotherapeutic agents in CRC, and this approach is readily available to enter clinical trials. From a translational point of view, there may be great variability in sensitivity to the inhibitors between individual patients.

The use of flow cytometry in heat-shock protein (HSP) research is increasing rapidly due to the high sensitivity and versatility of the technique. The method allows the simultaneous analysis of multiple proteins within numerous cell types in a heterogeneous sample, providing advantages over alternative techniques, such as ELISA and Western blotting. As a result, flow cytometry is becoming the leading technique used in this area of research. The current chapter describes the methodology for preparing samples for this technique and outlines two protocols for the analysis of surface- and intracellular-localised HSPs.

The enzyme-linked immunosorbent assay (ELISA) is an immunological technique which is used to determine the presence or quantity of an antigen within a sample. ELISAs rely on the use of at least one antibody (Ab) specific for the antigen being measured. This antibody is covalently linked to an enzyme which is detected through the use of an enzymatic substrate, which can be colorimetric, fluorogenic, or chemiluminescent. The ELISA for Hsp72 described here is a typical indirect sandwich ELISA, which can be used for measuring Hsp72 from cellular/tissue extracts, tissue culture supernatant, and serum. Typically, a 96-well ELISA plate is coated with a specific antibody which captures Hsp72 from the sample, and another antibody specific for a different Hsp72 epitope is used to detect Hsp72. An enzyme-labelled species-specific antibody conjugate is then applied which is consequently detected using a colorimetric enzyme substrate. The quantity of Hsp72 present in the samples is interpolated using a standard curve of known amounts of pure Hsp72.

Treatment of chronic lymphocytic leukemia (CLL) remains a challenge due to the frequency of drug resistance amongst patients. Improving the delivery of chemotherapeutic agents while reducing the expression of anti-apoptotic Heat Shock Proteins (HSPs) within the cancer cells may facilitate in overcoming this drug resistance. We demonstrate for the first time that sub-lethal doses of chemotherapeutic agents can be combined with membrane fluidizing treatments to produce a significant increase in drug efficacy and apoptosis in vitro. We show that fluidizers result in a transient decrease in intracellular HSPs, resulting in increased tumor-cell sensitivity and a membrane-associated induction of HSP gene expression., (2010 Elsevier Ireland Ltd. All rights reserved.)

Introduction: Monocytes/macrophages accumulate in the rheumatoid (RA) synovium where they play a central role in inflammation and joint destruction. Identification of molecules involved in their accumulation and differentiation is important to inform therapeutic strategies. This study investigated the expression and function of chemokine receptor CCR9 in the peripheral blood (PB) and synovium of RA, non-RA patients and healthy volunteers., Methods: CCR9 expression on PB monocytes/macrophages was analysed by flow cytometry and in synovium by immunofluorescence. Chemokine receptor CCR9 mRNA expression was examined in RA and non-RA synovium, monocytes/macrophages from PB and synovial fluid (SF) of RA patients and PB of healthy donors using the reverse transcription polymerase chain reaction (RT-PCR). Monocyte differentiation and chemotaxis to chemokine ligand 25 (CCL25)/TECK were used to study CCR9 function., Results: CCR9 was expressed by PB monocytes/macrophages in RA and healthy donors, and increased in RA. In RA and non-RA synovia, CCR9 co-localised with cluster of differentiation 14+ (CD14+) and cluster of differentiation 68+ (CD68+) macrophages, and was more abundant in RA synovium. CCR9 mRNA was detected in the synovia of all RA patients and in some non-RA controls, and monocytes/macrophages from PB and SF of RA and healthy controls. CCL25 was detected in RA and non-RA synovia where it co-localised with CD14+ and CD68+ cells. Tumour necrosis factor alpha (TNFα) increased CCR9 expression on human acute monocytic leukemia cell line THP-1 monocytic cells. CCL25 induced a stronger monocyte differentiation in RA compared to healthy donors. CCL25 induced significant chemotaxis of PB monocytes but not consistently among individuals., Conclusions: CCR9 expression by monocytes is increased in RA. CCL25 may be involved in the differentiation of monocytes to macrophages particularly in RA.

We report a kinetic study of the reaction between superoxide and an important intracellular form of vitamin B(12), cob(II)alamin. Superoxide is implicated in the pathophysiology of many inflammatory diseases, whereas vitamin B(12) derivatives are often beneficial in their treatment. We found that cob(II)alamin reacts with superoxide at rates approaching those of superoxide dismutase itself, suggesting a probable mechanism by which vitamin B(12) protects against chronic inflammation and modulates redox homeostasis.

The detection and confirmation of bloodstains as being human in origin is important in crime scene investigations. There are a number of blood detection methods currently available. The aim of this work was to develop an assay capable of detecting the presence of human blood from both liquid blood samples and dried bloodstains. A simple, direct competitive ELISA was developed utilising a polyclonal antibody against human IgG. Once optimised, the ELISA was found to be specific for human IgG, with no cross-reaction observed with pig, sheep, cow, goat, horse and rabbit IgG. The assay was also found to be sensitive, with a detection limit of 0.1 microg/mL. This compares favourably with leading blood detection methods. The assay was able to confirm the presence of human blood in blood mixtures, in stains on a variety of surfaces and also gave positive results with bloodstains that were up to 1 year old. The assay was simple to use, rapid and highly reproducible. The ELISA performance makes it suitable for development as a kit to rival currently used methods for the routine detection of human blood at crime scenes. Further applications of the anti-human IgG antibody are reported, including immunodot assays and a sandwich ELISA format. The methods described here are simple, reliable assays for the identification of human blood and are presented as viable alternatives to existing techniques for blood detection.

Oxidative stress is a feature of many chronic inflammatory diseases. Such diseases are associated with up-regulation of a vitamin B(12) (cobalamin) blood transport protein and its membrane receptor, suggesting a link between cobalamin and the cellular response to inflammation. The ability of cobalamin to regulate inflammatory cytokines suggests that it may have antioxidative properties. Here we show that cobalamins, including the novel thiolatocobalamins N-acetyl-l-cysteinylcobalamin and glutathionylcobalamin, are remarkably effective antioxidants in vitro. We also show that thiolatocobalamins have superior efficacy compared with other cobalamin forms, other cobalamins in combination with N-acetyl-l-cysteine (NAC) or glutathione (GSH), and NAC or GSH alone. Pretreatment of Sk-Hep-1 cells with thiolatocobalamins afforded robust protection (>90% cell survival) against exposure to 30 microM concentrations of the pro-oxidants homocysteine and hydrogen peroxide. The compounds inhibited intracellular peroxide production, maintained intracellular glutathione levels, and prevented apoptotic and necrotic cell death. Moreover, thiolatocobalamins are remarkably nontoxic in vitro at supraphysiological concentrations (>2 mM). Our results demonstrate that thiolatocobalamins act as powerful but benign antioxidants at pharmacological concentrations. Because inflammatory oxidative stress is a component of many conditions, including atherosclerosis, dementia, and trauma, their utility in treating such disorders merits further investigation.

Osteoprotegerin (OPG) is a major regulator of osteoclastogenesis, bone resorption and vascular calcification. OPG is produced by various cell types including mesenchymally derived cells, in particular, osteoblastic cells. Here we show OPG production by osteoblastic cells was stimulated by platelet-derived growth factor (PDGF) in two human osteosarcoma cell lines (MG63, Saos-2), a mouse pre-osteoblastic cell line (MC3T3-E1) and human bone marrow stromal cells (hMSC) by 152%, 197%, 113% and 45% respectively over 24 h. OPG was measured in the cell culture medium by immunoassay. PDGF isoforms AA, BB and AB show similar stimulation of OPG production. Message for OPG was also increased similarly to the increased secretion into the culture medium. Using specific inhibitors of cell signalling we demonstrate that PDGF acts through the PDGF receptor, PKC, PI3K, ERK and P38 and not via NF-kB or JNK. The importance of PDGF in fracture healing suggests a role for OPG production in countering bone resorption during the early phase of this process., ((c) 2008 Wiley-Liss, Inc.)

Heat shock proteins have been shown to be secreted from a number of cell types. Necrotic cells release heat shock proteins in a passive manner, whereas we, and others, have shown that viable cells secrete Hsp70 and Hsp60 through an active mechanism involving lysosomal vesicles and lipid rafts. This release of Hsp70 and Hsp60 is regulated, for example by being increased by elevated temperature. This article outlines procedures, using Hsp70 as the example, to: ensure the status of cells (viable, apoptotic or necrotic); identify the heat shock protein secreted; and quantify the secreted protein. Hsp70 has previously been quantified by ELISA, but newer methods are now being adopted, such as BIAcore and bead-based assays for use by FACS. These methods have the advantages of being more sensitive and requiring less sample than ELISA. The BIAcore has the potential to analyse Hsp70 ligands and provide affinity constants. The FACS bead assay system can be used to run multiplex assays.

There are an increasing number of studies reporting the presence of Hsps in human serum. We have investigated the release of Hsp70 into blood and culture medium from peripheral blood mononuclear cells (PBMCs), and whether this release is due to cell damage or active secretion from the cells. Intact Hsp70 was released from cells within whole blood and from purified PBMCs under normal culture conditions. Hsp70 release was rapid (0.1 ng/10(6) cells/h) over the first 2 h of culture and continued at a reduced rate up to 24 h (<0.025 ng/10(6) cells/h). Using viable cell counts and lactate dehydrogenase release we were able to confirm that the release of Hsp70 was not due to cellular damage. Hsp70 release was inhibited by monensin, methyl-beta-cyclodextrin, and methylamine, but not by brefeldin A. These data suggest that Hsp70 is released from cells via a non-classical pathway, possibly involving lysosomal lipid rafts.

Heat shock proteins (Hsps) are known to be induced in response to short-term stress. The present study aimed to evaluate the potential of Hsp70 as a biomarker of stress produced by increased temperature, osmotic pressure, and exposure to cadmium and sodium chloride in marine macroalgae and fresh water plant species. An indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) was developed with a working range of 0.025-10 microg ml(-1) using a monoclonal antibody raised against purified Hsp70 of Phaseolus aureus (mung bean). Fucus serratus (toothed wrack), Chondrus crispus (Stackhouse or Carrageen moss), Ulva lactuca (sea lettuce) and Lemna minor (common duckweed) sample extracts were stressed for up to 24 h and then tested in the IC-ELISA. The presence of Hsp70 and cross-reactivity of the monoclonal antibody was confirmed by Western blot. The heat shock response was confirmed in each species using a 2-h 42 degrees C treatment. Following heat shock, Hsp70 concentrations increased to a peak at 2 h (F. serratus) or 4 h (L. minor), after which concentrations decreased. Osmotic and cadmium stresses also resulted in elevated Hsp70 concentrations in samples of F. serratus and L. minor when compared with unstressed controls. In both, osmotic and metal stress, the production of Hsp70 increased to a maximum and subsequently decreased as the stressor levels increased. Results suggest that Hsp70 IC-ELISA could potentially be applied to the detection of stress in these aquatic species, although it would probably be most effective when used in conjunction with other measurements to provide a stressor-specific biomarker profile or fingerprint., (Copyright 2004 Taylor & Francis Ltd)

Background: Topical over-the-counter remedies exist to aid in the control of seborrheic dermatitis and chronic dandruff on a superficial level. Low-dose systemic oral nickel and bromide therapy has shown promise in providing improvement and eventual clearing of the disease., Objective: The purpose of this study was to further evaluate the effect of an orally administered low-dose, homeopathic mineral therapy (Potassium bromide 1X, Sodium bromide 2X, Nickel sulfate 3X, Sodium chloride 6X) on seborrheic dermatitis and chronic dandruff., Methods: Forty-one patients with seborrheic dermatitis and/or chronic dandruff were assigned to one of two treatment groups: Active (containing the medication) or placebo (vehicle). Study medication was administered in a placebo-controlled, randomly-selected, double-blind study for 10 weeks. At the end of 10 weeks all patients crossed over to the active medication, under a different label for an additional 10 weeks in an open study format., Results: Twenty-nine patients completed the 10-week blinded portion of the study. After 10 weeks of treatment, the disease state of the active patients improved significantly over that of the placebo patients (p<0.04). The placebo patients' condition before and after crossover to active treatment was also evaluated, showing significant improvement (p<0.01) 10 weeks after crossing over to active medication., Conclusion: Oral therapy using a low-dose homeopathic preparation combining Potassium bromide 1X, Sodium bromide 2X, Nickel sulfate 3X, and Sodium chloride 6X, provides significant improvement in seborrheic dermatitis and dandruff after 10 weeks of dosing.