Cell culture proliferation

EDI), and water to produce a group of biodegradable PU foams. The interconnected pores varied in size from 10 to 2 mm in diameter. Rabbit bone-marrow stromal cells cultured on the materials for up to 30 days formed multilayers of confluent cells and were phenotypically similar to those grown on tissue culture PS. It supported the adherence and proliferation of both bone-marrow stromal cells and chondrocytes in vitro. In subdermal implants the investigators found that the material showed infiltration of both vascular cells and connective tissue. 

The cell fusion mixture is transferred to a culture medium containing hypoxanthine, aminopterin and thymidine (HAT medium). Unflised myeloma cells are unable to grow as they lack HGPRT. Unflised normal spleen cells can grow but their proliferahon is limited and they eventually die out. The hybridoma cell can proliferate in the HAT medium as the normal spleen cell supplies the enzyme which enables the hybridoma to utilize extracellular hypoxanthine. 

To evaluate cell proliferation in the hydrogel, the L929 cells were immobilized at a density of 1.0 x 105 cells/mL. As a control sample, L929 cells were seeded onto a conventional cell culture plate at a density of 0.5 x 104 cells/mL. 

Cell Culture Substrates for Specific Cell Proliferation. 178... 

We wondered why NSCs proliferated exclusively on surfaces with EGF-His ligands anchored by coordination. We focused on two aspects in particular the conformational integrity of coordinated EGF-His and the stability of coordinate bonds at the interface. Conformational information was acquired with multiple internal reflection-infrared absorption spectroscopy (MIR-IRAS) [97]. The stability of coordinate bonds was assessed by culturing NSCs on a surface with a small region of EGF-His ligands anchored by coordination. This spatially restricted EGF-His anchoring enabled an intuitive exploration of EGF-His release under cell culture conditions. 

In order to reduce the time required to confirm the accumulation of a given recombinant protein, we have developed a cell culture system in which transgenic alfalfa callus material produced at the proliferation step of Agrobacterium-based transformation is used to initiate cell cultures. These cell suspensions can be subcultured to sustain batch production of modest protein amounts. The protein blot shown in Fig. 1.2 demonstrates our ability to detect a recombinant protein in total... 

Fig. 7.3.2. Effect of estradiol and BP-5 on MCF7 cell proliferation and gene expression. MCF7 cells were grown in estrogen-depleted medium. Estradiol or BP-5 were added to cultures and cell number (proliferation), protein determination (PgR and pS2), gene expression (pS2 mRNA) and luciferase expression were estimated at the appropriate times, as indicated in the Methods Section.

Based on the results from those cell calibration assays, the routine incubation period was set at 3 days. During that interval, some of the cells will proliferate, some will be quiescent but metabolically active, and some will die, which is the case for most primary cell cultures. Because these cells are cultured for periods far shorter than their typical population doubling time (about 6 to 7 days), these cells should not fully adapt to the tissue culture conditions and, therefore, maintain their primary cell phenotype. 

We have chosen animal cell cultures for that purpose, bearing in mind the following advantages of such system (i) the action of peptide results in a limited number of integral responses, when a variety of biochemical mechanisms gives rise to uniform effects, such as cell death or stimulation/inhibition of cell proliferation rate (ii) the test requires low, picomolar amounts of peptides (iii) the results are treated by simple and reliable statistic methods. 

Over 300 peptides isolated in our laboratory were studied in one or more tumor or normal cell cultures [39-44]. Part of the results obtained is summarized in Table 2.3. Over 75% of the peptides showed pronounced proliferative or antiproliferative activity in at least one cell type (Fig. 2.3). As a rule, tumor cells are more sensitive to peptide action. Besides the cell type, experimental conditions such as cell density or composition of the culture medium also affected the overall effect. In several cases (13%, Fig. 2.3) even the sign of the effect was peptide concentration dependent. Generally, experiments with cell cultures conform with the view that the main physiological function of cell and tissue peptidomes is control of long term processes and the homeostatic balance (i.e. cell differentiation, proliferation and elimination). The overall effect of peptide pools is achieved by concerted action of total sets of peptides rather than by single components. The molecular mechanisms of peptide action in cells requires concrete study in each individual case and are the subject of current research. 

The following basic protocols may be used alone or combined with other staining procedures in multiparameter flow cytometry experiments. Although they are illustrated with data from cells that proliferate in suspension, these protocols may be easily modified for the analysis of cells isolated from tissues or adherent cells in culture, by incorporating an initial step for the preparation of single cell suspensions. The assays are conducted at room temperature, unless otherwise noted. 

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