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Protein Secretion Systems in Microbial and Mammalian Cells
Published in Juan A. Asenjo, Separation Processes in Biotechnology, 2020
Gene transfer techniques, such as calcium phosphate precipitation and electroporation, permit the introduction of DNA into mamalian hosts. Three major types of vector have been developed, each allowing establishment of permanent cell lines producing desired proteins. The bovine papilloma virus (BPV) vector can be maintained stably as a multicopy extrachromosomal plasmid in the host (Lusky and Botchan, 1984). Retroviral vectors may be propagated in virus particles which can infect a wide range of host cell lines, and the DNA can integrate into the chromosomes (Cepko et al., 1984). The third type of vector also integrates into the chromosomes upon transfection, but it carrys a selectable marker (Wigler et al., 1977; Robins et al., 1981). The gene encoding the protein of interest is generally introduced on a separate similar vector by co-transfection. Selectable markers such as the thymidine kinase (TK) gene (Wigler et al., 1977) and the dihydrofolate reductase (DHFR) gene (Lee et al., 1981) have been used to complement TK- or DHFR-deficient hosts. In addition, dominant selectable markers such as the bacterial gpt (xanthineguanine phosphoribosyltransferase) gene (Mulligan and Berg, 1981) and the neo (neomycin phosphotransferase) gene (Southern and Berg, 1982) can be used for selection in a much wider range of cell lines. Both constitutive and regulated promoters have been used for the expression of desired genes.
Tracers
Published in Werner Käss, Tracing Technique in Geohydrology, 2018
The Environmental Agency at the Ministry for Environment, Nature Conservation, and Reactor Safety of the Federal Republic of Germany ordered basic toxicological investigations to be carried out for the most common fluorescent dyes. These investigations were conducted using the salmonella-microsomas test and with the cytogenetical analysis in the permanent cell line V 79 of the Chinese hamster. The test results indicated that the following substances can be used in groundwater tracing with no qualms as to their toxicity to humans: uranine, eosin, sulforhodamine B, amidorhodamine G, pyranine tinopal CBS-X, tinopal ABP liquid, and sodiumnaphthionate. With this, it is assumed that the injected substance amount is minimized for the investigation program. The dyes not listed here are being tested further (UBA 1997).
Evaluation of Food and Food Contaminants
Published in William J. Rea, Kalpana D. Patel, Reversibility of Chronic Disease and Hypersensitivity, Volume 5, 2017
William J. Rea, Kalpana D. Patel
Four DDT metabolites, p,p′-2,2-bis(chlorophenyl)-1-chloroethylene (DDMU), p,p′-2,2-bis(chlorophenyl)-1-chloroethane (DDMS), p,p′-2,2-bis(4-ch1oropheny1)acetonitrile (DDCN), and p,p′-2,2-bis(chlorophenyl)acetic acid (DDA), were selected based on their presence in environmental samples in Germany such as in sediments from the Mulde River and Teltow Canal. All assays used o,p′-DDT as reference. Cytotoxicity was measured by neutral red retention with the permanent cell line RTG-2 of rainbow trout (Oncorhynchus mykiss). Dioxin-like activity was determined using the 7-ethoxyresorufin-O-deethylase assay. The estrogenic potential was tested in a dot blot/RNAse protection assay with primary hepatocytes from male rainbow trout (O. mykiss) and in a yeast estrogen screen (YES) assay.
Spectral analysis and biological activity assessment of silver doped hydroxyapatite
Published in Journal of Asian Ceramic Societies, 2021
Umit Erdem, Busra Moran Bozer, Mustafa B. Turkoz, Aysegul U. Metin, Gurcan Yıldırım, Mustafa Turk, Saffet Nezir
For the test method, L929 fibroblast cells and osteoblast cells stained with neutral red were used. This method was used for testing the nonspecific cytotoxicity of leachable components belonging to the test substances after diffusion through agar or agarose (ISO 7405) using permanent cell lines red vital stain dye coated with an agar layer on the cell. All the cell lines (2 × 105 cells/well) were seeded into the sterile flat-bottomed 6-well plates (each well is 34.8 mm) containing DMEM with 10% FBS supplemented with 1% PS and %1 l-glutamine and incubated overnight at 37°C and 5% CO2-humidified atmosphere for 24 h. After discarding the medium, the agar medium was prepared on the cells for applying the samples. Here, the agar medium was composed of 50% agar and 50% DMEM allowed to gel at room temperature for 30 min. Neutral red was applied to the gelled agar medium, protected from light, and left for staining for 15 min. Neutral rejection was removed from the medium, and the extracts of 2, 5, and 10% Ag-doped HAp were applied to a concentration of 1/1 (100%) on the discs obtained from a 5 mm pre-filter and new systems were exposed to the incubation process for 24 h. In this experiment, the presence of leachable toxic substances is manifested by a change in the color of the dye (a type of membrane integrity test) during the lysis of cells (a type of membrane integrity test) as a result of the fact that both the concentrations of diffusing agents and cytotoxicity are high enough. Thus, it can be revealed that it is simple and inexpensive to use as a cytotoxicity determination method.
Monte Carlo parameter estimation and direct simulation of in vitro hyperthermia-chemotherapy experiment
Published in Numerical Heat Transfer, Part A: Applications, 2021
Nilton Pereira da Silva, Leonardo Antonio Bermeo Varon, José Mir Justino da Costa, Helcio Rangel Barreto Orlande
The hyperthermia treatment of cancer involves the heating of tumor cells in order to make them more susceptible to other kinds of treatment (like chemotherapy or radiotherapy) or to directly kill them with high temperatures and long exposures that cause permanent cell damage (in this last case, hyperthermia is usually referred as thermal ablation) [1–3]. The cytotoxic effect of hyperthermia combined with chemotherapy was examined by Tang and McGoron [4, 5] with in vitro experiments. They reported the positive adjuvant effects of thermal therapy on chemotherapy. Although the combined hyperthermia and chemotherapy present advantages for treating cancer, further studies need to be conducted on the synergistic use of heat and drugs [6]. Hyperthermia as a therapeutic technique is not applied only in oncology, but also in physiotherapy, urology, cardiology and ophthalmology [7].