Dye exclusion test

Cells are harvested from the dish by EDTA treatment, resuspended and rinsed in PBS-CMF, and finally resuspended at the desired concentration in serum free medium buffered at pH 7.2 with Hepes and filtered through 0.22 fim MiUipore membranes. Cell viability is checked by trypan blue dye exclusion test, and aggregates, if present, can be removed by filtration through cell strainers (Costar). 

Cell viability was determined using trypan blue dye exclusion test. 

Cell viability. Exponentially growing HL-60 cells, 15 x 106 in 75.0 mL of RPMI 1640, were incubated with either ATRA, or UDMA or BDDMA for five days. The total cell number was determined every day using the trypan blue dye exclusion test. Both the cellular proliferation - as area under curves (AUC) - and cellular mortality were calculated. 

It is well documented that normal human bronchial epithelial cells release a variety of cytokines and chemokines, and proinflammatory cytokines such as IL-la, IL-1 3, and TNF-a stimulate their production in vitro (Fig. 11) [61], We cultured human normal and transformed bronchial epithelial cells, and studied the effect of EM, CAM, and RXM on IL-6 [17], IL-8 [60, 62] and GM-CSF [63] production. Among the antimicrobes tested, only 14-membered-ring macrolides EM, CAM, and RXM showed an inhibitory action on cytokine release by unstimulated and cytokine-stimulated human bronchial epithelial cells (Fig. 12). In contrast, a 16-membered ring macrolide josamycin (JM) failed to show such effects. LDH release assay, a trypan blue dye exclusion test, and a colorimetric MTT assay showed that this effect was not due to cytotoxicity. Percent inhibition of IL-8 protein release in human primary bronchial epithelial cells was 25.0 5.67% at lO M, which is comparable with clinically observed serum concentration. 

Exposing HeLa S3 cells at 37 C to varied concentrations of, respectively, Fru-Trp (0.1 UM - 1 mM), NO-Fru-Trp (0.1 yM - 1 mM), and NaN02 (0.6 yM - 6 mM) for varied periods of time (1-36 hr) does neither affect their viability (trypan blue dye exclusion test) nor capability to synthesize RNA or protein but is of considerable Influence on DM synthesis in the case of NO-Fru-Trp and NaN02> but not in the case of Fru-Trp which continues to be ineffective. None of the three compounds tested is of significant influence on cell number. Both NO-Fru-Trp and NaN02 stimulate DNA synthesis a maximum of activity ([ H]thymidine incorporation) exists at the 24 hr time point of incubation, with NO-Fru-Trp, for instance, generating a 2.5-fold increase (over control) at 1 mM concentration in the medium while NaN02, at comparable concentration, Increases DNA synthesis by a factor of 1.6 over control. 

Platelet viability can be assayed easily by either a dye exclusion test or by measuring the membrane potential using a carbocyanine dye. Platelet activation can also be measured easily by measuring increase in cytoplasmic Ca concentration. 

Capillary shear tests were performed on low density (50 g fresh weight 1" ) suspensions of M. citrifolia using the apparatus illustrated in Fig. 1. Under both laminar and turbulent conditions [54,120,121], the relative viability of the suspension, evaluated using the Evan s Blue dye exclusion technique, was found to fall with exposure time in the loop. Loss of viability is well described by a first-order model ... 

When tested on cell counts, dye exclusion and fine structure of guinea-pig thymocytes, mercuric chloride in concentrations which inhibit DNA synthesis, most often caused cell degeneration, with condensed nuclear chromatin and disintegrating cytoplasmic organelles [170]. 

The antioxidant activity afforded by NO is most important if it translates into a meaningful therapeutic event. To test the importance of this reaction pathway to cell survival, we compared viability of the cells exposed to in the presence or absence of NO, When cells are exposed to 20 pM there was appreciable loss of cell membrane integrity as measured by trypan blue dye exclusion (Kelley et al, 1999) the addition of NO deaeased cell membrane damage (Figure 6). 

The prepared cell line is collected on the 7th day is washed, resuspended in complete medium containing GM-CSF and IL-4 and 2 mL and is plated in the wells of a six-well culture plate. lmL of test chemical also prepared in complete medium with GM-CSF and IL-4 is added to each well and the plate is incubated for 24 h. Viability of the DC after test chemical exposure is then assessed by propidium iodide dye exclusion using a Coulter Epics XL flow cytometer (Ryan et al. 2004). 

The pH value of dialyzed solutions was within the 7.0-7.4 range that corresponds to the normal pH of biological systems. Taking into account that the average thickness of cell membranes is about 8-10 nm, we can suppose that the synthesized nanoparticles ( 2 nm) can penetrate through the cell membranes. The particles can be used in native, solubilized and modified forms. The senthisyzed nanoparticles were tested on the rat hepatocyte cells using the standard procedure of trypan blue dye exclusion [7]. For a particle concentration up to 50 mg/1, the toxic effect has not been detected. 

SECM inside living cells is a very challenging task. For this, electrode with submicron size tip diameter, special low current potentiostat, and appropriate shielding are needed. The obtained SECM signal is highly valuable from point of view of cell physiology if the measurement causes only minor invasion. It was shown [138] by trypan blue exclusion tests that MCF-lOA breast cells can survive intracellular voltammetric experiments. The test is based on the dye out pumping property of the live cells [139]. That means if trypan blue dye is added (5 pM solution) to the immobilized cells, then after a few hours the dead cells appear blue but live cells appear uncolored. 

A need exists to fractionate the solutes in the dye filtrate into retained organic and passed inorganic salt fractions. The passage of simple electrolytes occurs through ultrafiltration membranes and ion-exclusion hyperfiltration membranes at a high salt concentration. Both types of dynamic membranes were tested. 

---