Overview alamarblue is a cell viability assay reagent which contains the cell permeable, nontoxic, and weakly fluorescent blue indicator dye called resazurin. Plant compounds are a potential source of new antimicrobial molecules against a variety of infections. To produce the 100% reduced form of alamarblue, simply autoclave a sample containing media and alamarblue. Plant extracts suppose complex phytochemical libraries that may be used for the. Assessment of the alamar blue assay for cellular growth and viability in vitro. Resazurin is nontoxic and will readily penetrate into viable cells whereupon it is metabolised. For equation 2, it is necessary to include the fluorescence value for alamarblue in its fully reduced form. Overview alamarblue can be used in a wide range of scientific research areas and applications including experiments involving cell proliferation, cell viability, bioassays for relative cytotoxicity, cytokine assays, cell metabolism studies, drug susceptibility, and toxicology studies simple and easy workflow just add the readytouse alamarblue solution to the cells, incubate for at least 1.
This change can be detected using fluorescence or absorbance measurement. General method for measuring cytotoxicity or proliferation using alamarblue. Ae markaki dept of engineering, university of cambridge 062009. Evaluation of the alamarblue assay for adherent cell. In this study, we determined the methodology for application of the assay to radiation response experiments in 96well plates. The bioassay may also be used to establish relative cytotoxicity of agents within various chemical classes 3. Section 5 fire fighting measures foam, dry powder, or carbon dioxide fire extinguisher as appropriate to local fire conditions. Resazurin cell viability assay offers a simple, rapid, reliable, sensitive, safe and costeffective measurement of cell viability. Microtiter plate reader for reading absorbance at 570nm and 600nm.
Staphylococcus aureus biofilms were treated with eleven. Colorimetric alamarblue assay as a bacterial concentration. An assay used to quantify the proliferation of various human and animal cell lines, bacteria and fungi, and assess relative cytotoxicity of agents within various chemical classes. Alamar blue ab is a watersoluble dye that has been previously used for quantifying in vitro viability of various cells fields and lancaster, 1993.
Colorimetric alamarblue assay as a bacterial concentration and spoilage index of marine foods. Safety data sheet section 1 chemical name and company details. Sizes available alamarblue can be used for applications such as functional assay, immunofluorescence, and elisa. We showed that flavonoltreated prostate cancer cells were stained less intensely with crystal violet than untreated cells at nontoxic concentrations. The readout options for the assay are absorbance or fluorescence. Alamarblue assay definition of alamarblue assay by medical. For each compound, one can determine the cytotoxic concentration exerting 50% cell damage, that is 50% decrease in the fluorescent signal of control cells. Measuring cytotoxicity or proliferation alamarblue assay. Comparison of alamar blue and mtt assays for high throughput screening. When incubated withviable cells, the reagent changes color from blue to red and becomes fluorescent figure 1. Format recommended incubation time bottomread fluorescence 10. A simple method to measure cell viability in proliferation. If however the spilled alamar blue has been in contact with a culture the spill should be. Disadvantages of metabolic reduction assays drug discovery.
Jain, tip biosystems pte ltd, singapore photopette cell makes the measurement of cell viability and proliferation easy and efficient. The mtt assay enables spectrophotometric measurement of mitochondrial dehydrogenase activity in. The alamarblue hs and alamarblue cell viability reagents are readytouse resazurinbased reagents that function as cell health indicators by using the reducing power of living cells to. You can create and edit multiple shopping carts edit mode allows you to edit or modify an existing requisition prior to submitting. Resazurin or gfafc was incubated with 10,000 du145 cells per well for 18 hr prior to measuring atp as an indicator of cell viability. The dye incorporates an oxidationreduction redox indicator that both fluoresces and change color in response to the chemical reduction of growth medium due to cell growth. You will have to determine the length of time for development though. Seven drugs, astemizole, daunorubicin, ellipticine, fluphenazine, terfenadine, thioridazine and trifluoperazine, had percent viability results of 55% or less in the alamar blue assay. The alamarblue assay was carried out as is described here in detail.
Cell viability and proliferation are important metrics of a material or construct for tissue repair. Microplate alamar blue assay for susceptibility testing of candida albicans biofilms. A staticcidal assay for trypanosoma brucei to aid hit. The performance of flow cytometry and the microplate alamar blue assay in determining susceptibility of mycobacterium tuberculosis was assessed by testing 150 brazilian isolates. Measure absorbance at wavelengths of 570 nm and 600 nm after required incubation. This is a trusted and established reagent which has been available since 1993. Cell viability assessment using the alamar blue assay.
Therefore, a lot of research on cell viability assay has been carried out. Measuring cytotoxicity or proliferation alamarblue assay protocol. Resazurin, the active ingredient of alamarblue reagent, is a nontoxic, cellpermeable compound that is blue. Description the celltiter blue cell viability assay provides a homogeneous. Bioteks visual abstracts are brief, animated presentations that describe the workflow of a single specific application. Multiple applications of alamar blue as an indicator of. Flavonoids reduce colorimetric dyes such as mtt and alamar blue in the absence of cells. In vitro antimycobacterial activity of six cameroonian medicinal plants using microplate alamarblue assay.
The simple protocol involves adding a single reagent directly to cells cultured in serumsupplemented medium. Upon entering living cells, resazurin is reduced to resorufin, a compound that is red in color and highly fluorescent. Microplate alamar blue assay how is microplate alamar blue. In addition, two blank wells media only and two controlwells media plus alamarblue are defined on the plate. Among many evaluation methods of cell viability, the alamar blue method is widely accepted for its simple operation and. Read a new alamar blue viability assay to rapidly quantify oligodendrocyte death, journal of neuroscience methods on deepdyve, the largest online rental service for scholarly research with thousands of academic publications available at your fingertips. It is used as an oxidationreduction indicator in cell viability assays for both aerobic and anaerobic respiration. Among many evaluation methods of cell viability, the alamar blue method is widely accepted for its simple operation. Pijnenborgthe use of alamar blue assay for quantitative. The attached pdf talks of proscons of different assays. As a service to our customers we are providing this early version of the manuscript.
The absorbance of test and control wells was read at 540 and 630 nm with a standard spectrophotometer. Overview although alamarblue is a simple and easy to use reagent for cell proliferation and viability experiments, there are some key facts and tips which will help you get the best results in your research optimizing your experiments using these tips will improve your assay accuracy to achieve better results using alamarblue. Multiple applications of alamar blue as an indicator of metabolic function and cellular health in cell viability bioassays. A staticcidal assay for trypanosoma brucei to aid hit prioritisation for progression into drug discovery programmes. Pettit, microplate alamar blue assay for susceptibility testing of. A malaria gametocytocidal assay using oxidoreduction. Comparison of flow cytometric and alamar blue tests with the. Resazurin, the active ingredient of alamarblue reagent, is a nontoxic, cellpermeable compound that is blue in color and virtually nonfluorescent. Resazurin reduction assays for screening of antitubercular. Performing viability assays on primary cells using trypan blue and aopi duration. Just dilute into your cell growth media and place on your cells. Due to the fact that it is extremely stable and more importantly nontoxic to the cells, continuous monitoring of cultures over time is possible ahmed et al. Xtt can be used to assay cell proliferation, cell viability, andor cytotoxicity.
The resazurin assay also known as alamar blue assay offers a simple, rapid, and sensitive measurement for the viability of mammalian cells and bacteria. When incubated withviable cells, the reagent changes color from blue. Application of a high throughput alamar blue biofilm. Use of the viability reagent prestoblue in comparison with. Assessment of cell proliferation with resazurinbased. Comparison of alamar blue and mtt assays for high through. To produce the 100% reduced form of alamarblue, simply autoclave a sample containing media and alamarblue for 15 min. Alamarblue cell viability assay reagent quantitatively measures the proliferation of mammalian cell lines, bacteria and fungi. Alamarblue assay for cell proliferation bmg labtech. Berringtonviability reagent, prestoblue, in comparison with other. It uses the indicator dye resazurin to measure the metabolic capacity of cellsan indicator of cell viability.
Microplate alamar blue assay for susceptibility testing of. Sep 10, 2012 compared to the ttc viability assay, the alamar blue assay was. This is a pdf file of an unedited manuscript that has been accepted for publication. Oct 01, 2003 read colorimetric alamarblue assay as a bacterial concentration and spoilage index of marine foods, food control on deepdyve, the largest online rental service for scholarly research with thousands of academic publications available at your fingertips. Analysis of cell viability by the alamarblue assay request pdf. Lee, acumen research laboratories pte ltd, singapore, and a.
The 96well microplate alamar blue assay maba allows for the quantitative determination of drug susceptibility against any strain of replicating mycobacterium tuberculosis to be completed within a week at minimal cost. A 96well plate containing the cells and the compounds to be tested is prepared using standard methods. The celltiterblue cell viability assay provides a homogeneous, fluorometric method for estimating the number of viable cells present in multiwell plates. Synthesis and anticancer activity of new flavonoid analogs. A novel onestep, highly sensitive fluorometric assay to evaluate cellmediated cytotoxicity. Create mode the default mode when you create a requisition and punchout to biorad. This application note demonstrates how to measure an alamarblue assay for assessment of cell proliferation using a bmg labtech plate reader. Sizes available alamarblue can be used for applications such as functional assay. Alamarblue was added and its reduction measured 7 hours later. Pdf multiple applications of alamar blue as an indicator of. Section 6 accidental release measures in general the affected area should be treated as any other non hazardous liquid spill.
A measurement of the absorbance would register the unreacted, resazurin. The assay is based on detection of metabolic activity through an oxidationreduction redox indicator, which both fluoresces and changes colour in. The alamar blue assay is based on enzymatic reduction of indicator dye by viable cells and serves as an effective tool for assessing cell proliferation and as a screening technique. Microplate alamar blue assay for susceptibility testing of candida albicans.
The use of alamar blue assay for quantitative analysis of. Plate configuration provides for samples to be run in duplicate. Overview alamarblue can be used in a wide range of scientific research areas and applications including experiments involving cell proliferation, cell viability, bioassays for relative cytotoxicity, cytokine assays, cell metabolism studies, drug susceptibility, and toxicology studies simple and easy workflow just add the readytouse alamarblue. Raz b, iten m, gretherbuhler y, kaminsky r, brun r 1997 the alamar blue assay. Cell viability assays such as cell titer blue and alamar blue rely on the reducing property of viable cells and are commonly used as a. Sep 12, 20 the alamarblue assay is based on fluorometric detection of metabolic mitochondrial activity of cells. Resazurin 7hydroxy3hphenoxazin3one 10oxide is a blue dye, itself weakly fluorescent until it is irreversibly reduced to the pink colored and highly red fluorescent resorufin. Download fulltext pdf the use of alamar blue assay for quantitative analysis of viability, migration and invasion of choriocarcinoma cells article pdf available in human. We use various livedead stains, clone expansion assays, mtt, trypan blue, and ldh to support our primary in house viability assay which is the alamar blue rezasurin assay. Pdf the use of alamar blue assay for quantitative analysis of.
Drugs with antioxidant properties can interfere with cell. The trypan blue exclusion assay was selected as a reliable alternative for measuring cell viability. Dont let the color get really pink or you will have saturated the assay. Blue and weakly fluorescent resorufin red and highly fluorescent the assay incorporates a reductionoxidation redox indicator that both fluoresces and undergoes colorimetric change in response to. The alamarblue hs and alamarblue cell viability reagents are readytouse resazurinbased reagents that function as cell health indicators by using the reducing power of living cells to quantitatively measure viability. Analysis of cell viability by the alamarblue assay. Depending on the target, membrane or mitochondrial damage, the best test for sds is the calceinam assay, whereas the alamar blue assay. You will be able to modify only the cart that you have punchedout to, and wont have access to any other carts. The mtt assay was less sensitive detecting an effect for daunorubicin and trifluoperazine. Absorbance values may be affected by the type of plate whether round or flat bottom and the plate manufacturer. This protocol describes a 96well format relative titering method for lentiviral stocks, based on transduction at low moi, selection for transduced cells with puromycin or blasticidin, varying by vectors, and a cell viability assay. This assay has excellent performance compared to other resazurinbased cell proliferation kits such as alamarblue, prestoblue, or celltiterblue.