A simple method to measure cell viability in proliferation and cytotoxicity assays

Abstract

Resazurin dye has been broadly used as indicator of cell viability in several types of assays for evaluation of the biocompatibility of medical and dental materials. Mitochondrial enzymes, as carriers of diaphorase activities, are probably responsible for the transference of electrons from NADPH + H+ to resazurin, which is reduced to resorufin. The level of reduction can be quantified by spectrophotometers since resazurin exhibits an absorption peak at 600 etam and resorufin at 570 etam wavelengths. However, the requirement of a spectrophotometer and specific filters for the quantification could be a barrier to many laboratories. Digital cameras containing red, green and blue filters, which allow the capture of red (600 to 700 etam) and green (500 to 600 etam) light wavelengths in ranges bordering on resazurin and resorufin absorption bands, could be used as an alternative method for the assessment of resazurin and resorufin concentrations. Thus, our aim was to develop a simple, cheap and precise method based on a digital CCD camera to measure the reduction of resazurin. We compared the capability of the CCD-based method to distinguish different concentrations of L929 and normal Human buccal fibroblast cell lines with that of a conventional microplate reader. The correlation was analyzed through the Pearson coefficient. The results showed a strong association between the measurements of the method developed here and those made with the microplate reader (r(2) = 0.996; p < 0.01) and with the cellular concentrations (r(2) = 0.965; p < 0.01). We concluded that the developed Colorimetric Quantification System based on CCD Images allowed rapid assessment of the cultured cell concentrations with simple equipment at a reduced cost.

Cell viability assessment using the Alamar blue assay: A comparison of 2D and 3D cell culture models

“…Over the past 50 years, the AB assay has been widely used in studies of cell viability and cytotoxicity for biological and environmental applications (Rampersad, 2012;Vega-Avila and Pugsley, 2011;White et al, 1996). The bioassay can also be used to establish the relative cytotoxicity of agents within various chemical classes (Bopp and Lettieri, 2008;Borra et al, 2009;Mikus and Steverding, 2000;Miret et al, 2006). Using the REDOX indicator resazurin (oxidised form), it is possible to spectrophotometrically measure the cellular proliferation.…”

Biocompatibility of Orthodontic Resins: <i>In vitro</i> Evaluation of Monomer Leaching and Cytotoxic Effects

“…Cell viability was evaluated by Alamar Blue after three different exposure periods (24, 48 and 72 h) because it is a non-toxic dye and this allows it to be used for continuous monitoring of cell viability (Borra et al, 2009). Our results showed that all the materials were cytotoxic and that uncured samples caused a major decrease in cell survival compared to cured ones.…”

“…To date many studies focused on their physical properties, such as shear bond strength (Finnema et al, 2010), with less emphasis on RBDM biocompatibility, despite the proximity of their application site to the periodontal tissues which may cause inflammation of gingiva, oral mucosa and alveolar bone (Borra et al, 2009). Different studies, dealing with RBDM biocompatibility, are based on in vitro tests because of the ability to control the environment and cellular responses (Jagdish et al, 2009;Franz et al, 2003;Malkoc et al, 2010).…”

“…Cytotoxicity of cured and uncured orthodontic composites were evaluated by Alamar Blue (AB) staining (Biosource Internation, Camarillo, CA) in order to measure the influence of tested materials on cell viability (Borra et al, 2009). Cells were seeded in the lower chambers of a 24 trans well tissue culture plate (Costar, Corning Incorporated, NY, USA) at 20×10 3 cells/well density in 2 mL of complete medium.…”

Design, synthesis, and evaluation of novel 2-phenylpropionic acid derivatives as dual COX inhibitory-antibacterial agents

“…MIC values (Table 2) were revealed by fluorometric measurements using resazurin solution 63,64 . Chloramphenicol and ketoconazole were used as standard drugs in the activity test.…”