Stock Preparation Research Articles

Abstract 3045: Investigating the influence of glycosylation using cellular models of neuroblastoma

Abstract Background: Neuroblastoma is a common childhood malignancy with approximately 800 new cases diagnosed each year in the United States. Treatment of neuroblastoma directly depends on patient risk categories and includes combinations of surgery, radiation therapy, chemotherapy, stem cell therapy, immunotherapy, and retinoid therapy. Various characteristics of cancer in general and some specific to neuroblastoma cell lines contribute to a difficulty in delivering clinically relevant concentrations of therapeutic agents to neuroblastoma masses. Amplification of the MYCN gene in certain neuroblastoma cell lines correlates with increased cellular surface glycosylation levels as well as a worse prognosis for patients. Overexpression of the MUC1 gene is also associated with increased glycosylation, specifically type-O, in various cancers, but the role of MUC1 in neuroblastoma has not been thoroughly defined. The objective of this study was to evaluate the influence of type-O glycosylation on therapeutic uptake in different neuroblastoma cell lines. Methods: The neuroblastoma cell line SK-N-DZ, a MYCN-amplified line, was compared to SK-N-SH, a non-MYCN-amplified line (negative control) and BT-20, a highly glycosylated breast cancer cell line (positive control). Benzyl-α-GalNAc, an O-glycosylation inhibitor, was introduced to each cell line. To determine glycosylation levels in the respective cell lines, fluorescence assays were performed using CD227 (anti-MUC1) antibody. In vitro cytotoxicity assays were performed to determine maximum non-toxic concentrations of the glycosylation inhibitor (benzyl-α-GalNAc) and antibody (CD227) and to subsequently evaluate the functional role of glycosylation using cellular models of neuroblastoma. Results: The maximum non-toxic concentrations per 10,000 cells for each neuroblastoma cell line was 0.05 mg/ml of benzyl-α-GalNAc, and 6 µl of CD227 stock preparation. Preliminary results from fluorescence detection studies showed an increase in CD227-FITC antibody uptake by the MYCN-amplified neuroblastoma cells (SK-N-DZ) following exposure to the O-glycosylation inhibitor (benzyl-α-GalNAc). Cellular uptake of CD227-FITC by non-MYCN-amplified cell line (SK-N-SH) was less by comparison. Conclusion: Preliminary findings suggest that O-glycosylation is a barrier capable of limiting cellular uptake of agents in MYCN-amplified neuroblastoma cell lines. Ongoing studies are focused on investigating the influence of glycosylation inhibition on cytotoxicity of FDA-approved therapeutic agents. Citation Format: Meghan Cook, Alma Garcia, Katie Konieczny, Meghan Ferguson, Robert Campbell. Investigating the influence of glycosylation using cellular models of neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3045.

The effects of vegetable pickling conditions on the dynamics of microbiota and metabolites.

BackgroundSalting is a traditional procedure for producing pickled vegetables. Salting can be used as a pretreatment, for safe lactic acid fermentation and for salt stock preparation. This study aimed to provide valuable knowledge to improve pickle production by investigating the dynamics of microbiota and metabolites during the pretreatment and salt stock preparation processes, which have previously been overlooked. The differences in these process conditions would be expected to change the microbiota and consequently influence the content of metabolites in pickles.MethodsSamples, collected from eight commercial pickle manufacturers in Japan, consisted of the initial raw materials, pickled vegetables and used brine. The microbiota were analyzed by 16S rRNA sequencing and the metabolites quantified by liquid chromatograph-mass spectrometry. Statistical analyses helped to identify any significant differences between samples from the initial raw materials, pretreatment process and salt stock preparation process groups.ResultsUnder pretreatment conditions, aerobic and facultative anaerobic bacteria were predominant, including Vibrio, a potentially undesirable genus for pickle production. Under salt stock preparation conditions, the presence of halophilic bacteria, Halanaerobium, suggested their involvement in the increase in pyruvate derivatives such as branched-chain amino acids (BCAA). PICRUSt analysis indicated that the enhanced production of BCAA in salt stock was caused not by quantitative but by qualitative differences in the biosynthetic pathway of BCAA in the microbiota.ConclusionThe differences in the microbiota between pretreatment and previously studied lactic acid fermentation processes emphasized the importance of anaerobic conditions and low pH under moderate salinity conditions for assuring safe pickle production. The results from the salt stock preparation process suggested that the Halanaerobium present may provide a key enzyme in the BCAA biosynthetic pathway which prefers NADH as a coenzyme. This feature can enhance BCAA production under anaerobic conditions where NADH is in excess. The effects shown in this study will be important for adjusting pickling conditions by changing the abundance of bacteria to improve the quality of pickled vegetables.

SARS-CoV-2 variants reveal features critical for replication in primary human cells.

Since entering the human population, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2; the causative agent of Coronavirus Disease 2019 [COVID-19]) has spread worldwide, causing >100 million infections and >2 million deaths. While large-scale sequencing efforts have identified numerous genetic variants in SARS-CoV-2 during its circulation, it remains largely unclear whether many of these changes impact adaptation, replication, or transmission of the virus. Here, we characterized 14 different low-passage replication-competent human SARS-CoV-2 isolates representing all major European clades observed during the first pandemic wave in early 2020. By integrating viral sequencing data from patient material, virus stocks, and passaging experiments, together with kinetic virus replication data from nonhuman Vero-CCL81 cells and primary differentiated human bronchial epithelial cells (BEpCs), we observed several SARS-CoV-2 features that associate with distinct phenotypes. Notably, naturally occurring variants in Orf3a (Q57H) and nsp2 (T85I) were associated with poor replication in Vero-CCL81 cells but not in BEpCs, while SARS-CoV-2 isolates expressing the Spike D614G variant generally exhibited enhanced replication abilities in BEpCs. Strikingly, low-passage Vero-derived stock preparation of 3 SARS-CoV-2 isolates selected for substitutions at positions 5/6 of E and were highly attenuated in BEpCs, revealing a key cell-specific function to this region. Rare isolate-specific deletions were also observed in the Spike furin cleavage site during Vero-CCL81 passage, but these were rapidly selected against in BEpCs, underscoring the importance of this site for SARS-CoV-2 replication in primary human cells. Overall, our study uncovers sequence features in SARS-CoV-2 variants that determine cell-specific replication and highlights the need to monitor SARS-CoV-2 stocks carefully when phenotyping newly emerging variants or potential variants of concern.

Long-term physicochemical stability of acyclovir 5 mg/mL solution stored in polypropylene bags as a simulated hospital stock preparation.

To investigate the long-term chemical and physical stability of 5-mg/mL acyclovir solution in polypropylene bags stored at 5°C ± 3°C for 2 months in order to determine the feasibility of batch production by a centralized intravenous additive service. Eight empty 100-mL polypropylene bags (bags A) and 8 empty 250-mL bags (bags B) were respectively filled with 60 mL and 200 mL of 5-mg/mL acyclovir and 0.9% sodium chloride injection (NaCl) under aseptic conditions through a semiautomated manufacturing process and vacuum packed before storage at 5°C ± 3°C. Four bags A and 4 bags B were tested for chemical stability via a stability-indicating high-performance liquid chromatography (HPLC) method immediately after preparation (time 0) and after 7, 14, 21, 28, 35, 42, and 63 days. Samples for microbiological assay were collected on days 0 and 63 from 4 bags A and 4 bags B immediately after breaking the vacuum. Osmolality, pH, and physical stability were assessed by visual examination, Subvisible particle counting was performed on 6 additional bags (3 each of bags A and B). Mean percentage loss of acyclovir relative to the mean experimental concentration at time 0 was below 5% over the 63-day study period.. No significant differences of pH, no change in color and no precipitate were observed during the study. Subvisible particle counts were compliant with European Pharmacopoeia requirements. Acyclovir solutions remained sterile over the 63 days of the study. Extemporaneously prepared acyclovir 5 mg/mL solutions in 0.9% NaCl stored in polypropylene bags were chemically and physically stable over 63 days when stored at 5°C ± 3°C.

Processes and problems of pulp and paper industry: an overview

Abstract The pulp and paper industry is a highly energy-intensive and water-consuming industry. This industry is known for the utilization of a wide range of raw materials, containing cellulose fibers (generally wood, recycled paper, and agricultural residues), for the production of various grades of paper. There are several processes involved in the conversion of raw materials to the paper product such as raw material preparation, pulping, pulp washing and screening, bleaching, stock preparation, papermaking and chemical recovery. All the processes are facing issues regarding process efficiency, product quality, energy & water consumption, and cost and environment. There is a need for further improvement and upgrading the technologies but the scale of operations, technological obsolesce and cost of implementing new technologies are some of the major issues. The main thrust areas of pulp and paper processing require major interventions in the adoption of green and clean technologies.

Provision of the Continuum of Care to Noncommunicable Diseases Post-Floods in Kerala, India 2018.

Integrating noncommunicable disease (NCD) in health care delivery during emergency response posed a major challenge post-floods in Kerala. Kerala experienced an abnormally high rainfall during mid-2018 where more than 400 people lost their lives. State health officials and the Disaster Response Team were sensitized about the importance of including NCDs in the response action. More than 80% of patients with hypertension and diabetes were not under control in Kerala. Under the state NCD cell, an NCD expert group was consulted for drafting the treatment and referral strategies. Steps to tackle NCDs during the disaster response were formulated. The state NCD cell decided to integrate NCDs in the response measures. The technical guidance document by the World Health Organization South-East Asia Region was consulted to formulate actions. The activities were implemented in 6 steps: prioritizing of major NCDS, patient estimation and drug stock preparation, standard treatment protocol, mapping of referral facilities, public engagement, and daily reporting of NCD consultations. Prioritizing the continuum of care of NCDs during floods among the program managers and care providers was crucial. The health education and communication campaign was done to sensitize the known NCD patients to seek early care. Daily reporting of consultations was established.

Process audit for Stock Preparation System

Process audit for Stock Preparation System

The Keenest Stock Preparation Technologies for Board Paper

原料及び燃料価格の上昇に伴い，製紙工場では低廉古紙を使用し生産コストを低減したい意向がある。しかしながら低廉古紙には，現在使用している古紙よりも雑多な異物，難離解性古紙が含まれている。これらの異物や難離解性古紙は機器操業の安定を阻害し，品質に於いても重大な欠陥を引き起こす場合がある。

Experimental Evolution of Antifungal Resistance in Cryptococcus neoformans.

Cryptococcus neoformans, an opportunistic yeast-like fungal pathogen, has demonstrated resistance to all major classes of antifungals used to treat cryptococcal meningitis. However, combatting this fungal disease is an ongoing challenge among clinicians due to the evolution of antifungal-resistant strains. The limited availability of clinically approved antifungals has heightened the urgency to investigate the molecular mechanisms underscoring resistance. Studying how a fungal pathogen evolves to an antifungal drug in vitro using experimental evolution provides a simple, yet powerful approach to study the mechanisms of antifungal resistance. Experimental evolution involves the serial passaging of microbial populations under laboratory conditions, such that adaptive mutations can occur and be monitored in real time. This technique plays a key role in investigating the mechanisms of antifungal resistance in C. neoformans, and this can help in developing novel strategies to combat the emergence of resistance. Here, we outline how to make overnight cultures of C. neoformans and how to perform experimental evolution, and we present a spectrophotometric analysis to evaluate the evolution of antifungal resistance. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Growth and sample preparation of Cryptococcus neoformans Basic Protocol 2: Experimental evolution of antifungal resistance Basic Protocol 3: Analyzing the evolution of antifungal resistance Basic Protocol 4: Glycerol stock preparation.

Impact of embossing on liquid absorption of toilet tissue papers

Absorption capacity is a key feature of toilet tissue papers. Several parameters can affect their final absorption capacities, such as pulp composition, stock preparation, number of sheets, additives, bulk, grammage, and converting process parameters, such as the embossing operation. In this work, the absorption capacities of four different 2-ply industrial toilet tissue papers, as well as the respective base papers from the mother-reel was compared using the immersion method according to ISO 12625-8 (2010). Previously, these samples were characterized in terms of morphology, grammage, thickness, and bulk. It was concluded that the embossing operation noticeably increased the thickness and bulk of toilet tissue paper. Furthermore, it was also verified that among the various toilet tissue paper samples there was not a noticeable variation in the time of water absorption because the samples revealed similar morphology and porosity. However, it was found that the bulk increased more than 150%, resulting in an increase of water absorption capacity over 60%.

Feasibility and reliability of DEXA analysis after total ankle arthroplasty: A cadaver study

BackgroundAlthough the outcomes of total ankle arthroplasty (TAA) have improved, unsolved problems such as stress shielding remain. Although dual energy X-ray absorptiometry (DEXA) is the “gold standard” for evaluation of these issues, it is rarely used in patients after TAA. This study aimed to establish a scan technique and to assess the mechanical changes in bone density caused by bone stock preparation. MethodsEight fresh-frozen cadaver legs were investigated by DEXA before TAA, with implant in situ, and after implant removal. Scan surface, bone mineral content, and bone mineral density were analysed to assess mechanical bone mass changes. ResultsWe examined data for density changes by bone compression after TAA, and found “good” results for internal reliability but only “acceptable” results for external reliability. ConclusionsThe results were reliable and reproducible. Using the present data, mechanical and biological processes can be considered together to understand the postoperative phases of bone remodelling after TAA.

Introduction of Cooperative Process Optimization between Paper Machines and Stock Preparation

Introduction of Cooperative Process Optimization between Paper Machines and Stock Preparation

The Keenest Stock Preparation Technologies for Board Paper

The Keenest Stock Preparation Technologies for Board Paper

The Keenest Stock Preparation Technologies for Board Paper

The Keenest Stock Preparation Technologies for Board Paper

Stock Preparation Equipment for Household Paper

Stock Preparation Equipment for Household Paper

Preparation of Margarine Stock Rich in Naturally Bioactive Components by Enzymatic Interesterification.

Fully hydrogenated expanded press soybean oil (FHEPSO) rich in naturally bioactive components was prepared using Palladium on Carbon (Pd/C) catalyst. Interesterified fat was prepared from binary blends of FHEPSO and cold press corn oil (CPCO) with FHEPSO/CPCO mass ratios of 50:50, 40:60 and 30:70. Lipozyme RM IM (6 wt% of total substrate) was used in a supercritical CO2 system to catalyze the transesterification. The fatty acid compositions had no significant changes in the fats before and after interesterification, and trans-fatty acid (TFA) was not detected. The fatty acid compositions within triacylglycerol (TAG) were rearranged, and the amounts of trisaturated and triunsaturated TAG decreased, whereas that of mixed TAG increased as a result of interesterification. The enzymatic interesterified fats (EIEF) had a lower solid fat content (SFC), broader melting and plasticity ranges compared to the noninteresterified blend (NIB). According to X-ray diffraction (XRD), the predominant crystal form had changed from β to β'. EIEF contained 0.33-0.51 g/100 g phospholipids, 88.6-105.6 mg/100 g total tocopherols, and 916-1053 mg/100 g total phytosterols, which could confer health benefits. The results indicated that EIEF may have a potential use in trans-free margarine stock preparation.

Effect of Temperature and Time on Free amino acid profile in Thai chicken bone soup stock preparation

Effect of Temperature and Time on Free amino acid profile in Thai chicken bone soup stock preparation

A modified protocol for highly efficient EBV-mediated immortalization of human B lymphocytes from small volumes of peripheral blood serum

BackgroundMany human molecular and genetic studies require the use of a renewable biological material. Although primary fibroblast cell lines can be used for this purpose, there are disadvantages associated with human biopsies including the limited number of cell divisions. Peripheral blood has the advantage of being easier to obtain but also the drawback that blood cells produce only short-term cultures. Epstein-Barr virus is capable of transforming human B lymphocytes into indefinitely proliferating cells that can be maintained in tissue culture. Here, we report a convenient method of B-lymphocyte immortalization using small volumes of freshly collected human blood serum saturated with nucleated blood cells. Aim of the studyThe aim of the present study is modification and improvement of the protocol for highly efficient immortalization of human B lymphocytes from small volume of blood samples. Material and methodsCell line B95-8 was used as Epstein-Barr virus source for viral stock preparation. Immortalizing medium contains RPMI-1640, viral stock and additives. No feeder layer was used. ResultsAs result we present high efficient method for B lymphocytes immortalization with start blood volume less than 5 ml. ConclusionThe method is applicable for immortalization of B lymphocytes from small blood samples and is essential for studies involving children or patients when large blood volume sampling is impossible.

Hot Runner Mold Design of Fan Diverter Parts

In this study, we discuss the case of plastic parts for the production of fan steering gear shaft parts injection molding, and use POM plastic steel to produce plastic parts from traditional cold runners. Because of the parts have a hole, which need side slide. The runner produce more waste after plastic parts injection make the runner waste account for the cost is relatively high, the cost of stock preparation is relatively increased when the product quantity demanded is great. After the crushing treatment of the waste, the backfill will affect the quality, and in the crushing process, the volume generated will make the operator to withstand up to 130 dB of noise. The actual test results show that the production cycle reduce 6.25%, while the production yield increase by about 5% and material costs reduced by 2% . It can be recovered within a year, not to mention the increase of the quality and reduction the noise on the staff of the benefit is impossible to estimate.

The Adeno-associated Virus - A Safe and Promising Vehicle for Liverspecific Gene Therapy of Inherited and Non-inherited Disorders.

The first human adeno-associated virus (AAV) was originally discovered in 1960s as a contaminant of adenovirus stock preparation and thus it had not been of medical interest. Throughout the last three decades AAV has gained popularity to be used in gene therapy, mainly due to its replicative defectiveness and lack of pathogenicity in human. In addition, its ability to mediate stable and long-term expression in both non-dividing and dividing cells with specific tissue tropism makes AAV one of the most promising candidates for therapeutic gene transfer to treat many inherited as well as non-inherited disorders. Moreover, the use of AAV is not only restricted to overexpression of recombinant transgene, but also to over-express short hairpin RNA and microRNA to knockdown the expression of genes in targeted tissues. This review is organized into four parts. In the first part of the review, we discuss about the discovery and history of AAV, followed by detailed AAV biology such as virus genome, virus structure and its life cycle. In the second part of the review, the discussion is centred on the molecular mechanisms of AAV and tissue transduction, including receptor recognition and cell binding, endosomal entry, virus uncoating, nuclear entry and genome replication. Advantages and limitations of using AAV as a safe vehicle for gene delivery is also discussed. In the third part of the review, we discuss about the most commonly used AAV serotypes and variants isolated from human and non-human primates, focusing on their diverse tissue tropisms, transduction efficiency, immunological profiles and their applications in animal studies. Final part of the review focuses on the recent progress of in-vivo gene transfer using AAV for inherited and non-inherited diseases in both preclinical and clinical settings with a special emphasis on potential clinical applications of AAV in the field of liver disease.