Cell culture viability

Srinivasan, A., Hollinger, J.O., and Matyjaszewski, K. (2008) Synthesis, characterization, and in vitro cell culture viability of degradable poly(N isopropylacrylamide-co-5,6-benzo-2-methylene-l,3-dioxepane)-based polymers and crosslinked gels. /. Biomed. Mater. Res. A, 87A (2), 345. 

In the dialyzed batch start-up phase and the subsequent continuous operation a substantial increase in viable cell density and monoclonal antibody (MAb) titer was observed compared to a conventional suspension culture. The raw data, profiles of the viable cell density, viability and monoclonal antibody titer during the batch start-up and the continuous operation with a dialysis flow rate of 5 L/d are shown in Figures 17.6 and 17.7. The raw data are also available in tabular form in the corresponding input file for the FORTRAN program on data smoothing for short cut methods provided with the enclosed CD. 

All of these characteristics can be under the regulation of the cell and influenced by the cell culture conditions. The age of the cell monolayer in culture can have a profound impact on the quality of the barrier. In monolayers with actively dividing cells, resistance increases with time in culture as tight junctions form (see Fig. 15, Section III.C.4). Resistance reaches a plateau, then decreases as cell viability declines (Section III.C.4). Time in culture may also be a factor in the expression of polarity, which is related to tight junction formation as well as the state of differentiation of the cells (e.g., differential gene expression). 

In an attempt to mimic the physiological ratio of collagen type II to hyaluronan in the healthy human NP, Calderon et al. constructed hydrogels scaffolds composed of these two elements in a 9 1 (w/w) ratio [104]. Scaffolds were crosslinked with various concentrations of EDC/NHS, but it was found that 8 mM EDC/NHS resulted in a confined compressive modulus on the order of the native NP, while allowing for optimal rat mesenchymal stem cell (rMSC) viability and proliferation. Additionally, real time PCR results from rMSCs seeded on the scaffolds for 21 days indicated that the scaffolds promoted increased aggrecan expression and inhibited collagen type I expression compared to rMSCs cultured on monolayers. 

Table 7.1 Absence of C /polyvinylpyrrolidonc (C60/PVP) complex influence (5mg/ml) on cellular viability in cell culture MA-104. Endpoint - resazurin reduction...

A sufficient plating density has also to be chosen in order to obtain confluent monolayers within the limited time of viability of primary cell cultures. 

The hrefly Inciferin system is very sensitive and can be conpled to any enzymatic reaction that prodnces or nses ATP. For example, creatine phosphokinase can be determined by this method and hence be nsed in the diagnosis of myocardial infarction and mnscle disorders. The creatine phosphokinase converts AMP into ATP which then nndergoes the reaction with Inciferin as shown in Fignre 3.25. ATP pro-dnction is essential for every known life form and the firefly Inciferin system can be nsed to check for microbial life. Hence systems have been developed that use a portable luminescence workstation to monitor sanitation in food manufacturing and to check for sterile environments in technological workplaces. The system can also be applied in checking cell viability, for instance in cell cultures and to measure the toxic effects of chemicals on cells. 

If dechorionated embryos are cultured, some background reduction in viability and dysmorphology may be avoided by using multiwall plates that are not tissue culture treated. Tissue culture treated plates are designed to promote cell adhesion in cell culture, but this may cause the early stage zebrafish embryo to adhere to the plate with detrimental consequences. 

Acetaldehyde, benzene, butyraldehyde, iso-prene, styrene, and toluene in mouse lymphocytes cell culture for 3 hours produced no effect on either viability or proliferation. Lormaldehyde, catechol, acrylonitrile, propionaldehyde, and hydroquinone significantly inhibited T-lymphocyte and B-lymphocyte proliferation, inhibitory concentration (IC )jo 1.19 x 10" M to 8.20 x 10" M. Acrolein and crotonaldehyde inhibited T-cell and B-cell proliferation and acted on viability with ICjp 2.06 x 10 M to 4.26 X 10" M. Mixtures of acrolein, formaldehyde, and propionaldehyde or crotonaldehyde interactive effects at 0.5 and 1 x ICjo were observed " . 

As it is imperative that the plant-derived hiopharmaceutical product must be obtained repeatedly and on a consistent basis, a master cell culture bank, seed bank for transgenic plants, or virus seed stock for transient expression systems must be constantly maintained. Storage conditions must therefore he optimized to prevent contamination and ensure viability. Both transgene stability (e.g., reversion to wild type or sequence drift of plant virus expression vectors) and protein expression levels must be monitored in a representative plant of a given bank or stock to minimize any possible variation in expression levels that may affect safety and consistency of the hnal product. A program that monitors lot-to-lot consistency of the hiochemical and biological properties by comparing the product with appropriate in-house reference standards could he implemented as a fundamental component of product development. 

Grenier M, Sun P, Drobitch R. Effect of St. John s wort preparations on cell viability, induction of nitric oxide and ethoxyresomfin O-deethylase activity in glial cell culture. Proceedings of the 5th Annual S5Tnposium on Pharmaceutical Sciences. J Pharm Pharmaceut Sci 2002 5 106. 

Viability Viability of MSCs can easily be determined immediately after trypsin-ization via trypan blue or 7-amino-actinomycin D (7-AAD) exclusion. According to the specifications developed from our cell culture studies, viability should be >90%. In selected exceptional cases a lower limit of 70% viability of total harvested cells may be acceptable. 

The level of enzyme needed can influence the choice of preparation used for the study. Microsomal preparations from cell cultures allow the use of higher concentrations of active enzyme per unit volume than use of whole cells or cell lysates. The use of whole, viable cells allows the use of longer incubation times but at a lower enzyme concentration per unit volume. In addition, adequate oxygen transfer and nutrient concentrations are needed to maintain culture viability. These requirements impose limitations on cell concentration. In addition, microsomes cannot be efficiently prepared from all cultured cell types. We have found that standard microsome preparation procedures as used for human or rodent liver were unsuitable for isolating active enzymes from human lymphoblasts, and this appears to be a general property of cultured cell lines. Specific catalytic activities in microsomes were lower than for whole cell lysates. This loss of activity appears to happen in other mammalian cell systems which has led to the common use of whole cell lysates.With human lymphoblasts, shortening the length of... 

Trudell and our group, with the generous support of Dr. Hugo Jaurequi, conducted certain cell-culturing experiments on human hepatic cells. Since the effort was self-funded, the scope of the experiments was limited. Nevertheless, we felt we confirmed the viability of a specially designed polyurethane formulation as a substrate for cell growth. 

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