Description alamarbluetm fluorometric cell viability assay kit offers a simple, rapid, reliable, sensitive, safe and costeffective measurement of cell viability. The alamarblue hs and alamarblue cell viability reagents are readytouse resazurinbased reagents that function as cell health indicators by using the reducing power. Alamarbluetm detects cell viability by utilizing a blue and nonfluorescent dye resazurin, which is converted to a pink and fluorescent dye resorufin. Alphalisa assay development guide harvard university. Celltiterblue cell viability assay technical bulletin tb317. Monitoring viability using cellular reducing environment. 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. Allows your choice of assay format and method of detection. The goal was to make the biofilm susceptibility assay as similar to the nccls planktonic susceptibility assay 17 as possible.
Pdf multiple applications of alamar blue as an indicator of. Resazurin or gfafc was incubated with 10,000 du145 cells per well for 18 hr prior to measuring atp as an indicator. Performing viability assays on primary cells using trypan blue and aopi duration. Send us your target cell line, or use one of ours, and well take care of the rest. The celltiterblue assay is based on the ability of living cells to convert a redox dye resazurin into a fluorescent end product resorufin. Here, we optimized the standard alamarblue proliferationviability protocol for tumor spheroid cultures to enhance assay precision during. Singlestep, homogeneous, highthroughput cell quantitation. The alamarblue dye is a redox indicator that yields a colorimetric change and a fluorescent signal in response to metabolic activity. Our customizable cdc assays accurately detect cell lysis based on alamar blue staining of remaining live cells, using human serum complement as a probe.
Alamarblue cell viability reagent from thermo fisher. Transfer 100 l aliquots of each sample to replicate wells of a black 96well plate. Alamar blue ab is a watersoluble dye that has been previously used for quantifying in vitro viability of various cells fields and lancaster, 1993. The microdilution method circumvents some of the problems encountered by the. Tox8 store at 28ec this kit is designed for fluorometrically or spectrophotometrically determining cell number as a function of metabolic activity using the dye resazurin. This is a trusted and established reagent which has been available since 1993. The internal environment of proliferating cells is more reduced than that of nonproliferating cells. Alamarblue assay definition of alamarblue assay by. Resazurin is dark blue in color and has little intrinsic fluorescence. The alamarblue assay is designed to measure quantitatively the proliferation of various human and animal cell lines, bacteria and fungi. Living cells are metabolically active and are able to reduce via mitochondrial reductase, the nonfluorescent dye resazurin to the stronglyfluorescent dye resorufin fig.
Multiple applications of alamar blue as an indicator of metabolic. The alamar blue assay was used to measure the anoikisresistant cellspecific cytotoxicity of these compounds after 72 h of incubation. Alamar blue ab was diluted at 10% in media and then added to the well 100. Microtitre panel containing a range of antifungal drugs and alamar blue. In vitro toxicology assay kit resazurin based stock no.
Assessment of the alamar blue assay for cellular growth and viability in vitro. This product is however used in conjunction with fungal cultures and any hazards associated. Sample files alamarblue a november 15, 2012 download related products. Assessment of a simple, nontoxic alamar blue cell survival assay to monitor tomato cell viability. It is a nontoxic, water soluble, redoxsensitive dye that changes from its bluenonfluorescent state to a pinkhighlyfluorescent state upon reduction by viable cells. Alamar blue monitors the reducing environment of the living cell. The microplate was measured using the settings below. 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. The celltiter blue assay is based on the ability of living cells to convert a redox dye resazurin into a fluorescent end product resorufin. Measure fluorescence at 590 nm, using an excitation wavelength of 530560 nm. This assay has excellent performance compared to other resazurinbased cell proliferation kits such as alamarblue, prestoblue, or celltiterblue. Plate cells at approximately 104 cellswell in 96 well plates 10ml of 105. Analysis of cell viability by the alamarblue assay. 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.
Fluorescence can be read using 544 nm excitation and 590 nm emission wavelengths, or absorbance can be read using a spectrophotometer set at 570 nm. Upon entering living cells, resazurin is reduced to resorufin, a compound that is red in color and highly fluorescent. The bioassay may also be used to establish relative. For an alamar blue assay cell viablity experiment, how do i explain or interpret the results of a absorbance vs. Dont let the color get really pink or you will have saturated the assay. You will have to determine the length of time for development though. The fluorescence output is proportional to the number of viable.
The bioassay may also be used to establish relative cytotoxicity of agents within various chemical classes 3. Ab assay is a simple and reliable method for quantifying trophoblast. Complement dependent specific lysis of complement dependent. So one ends up going round and round without any clear conclusion. The alamarblue assay is designed to measure quantitatively the proliferation of various human. Comparison of alamar blue and mtt assays for high through. Resazurin 7hydroxy3hphenoxazin3one 10oxide is a phenoxazine dye that is weakly fluorescent, nontoxic, cellpermeable, and redox. A variety of parameters for the ab assay were standardized at the outset using s. Among many evaluation methods of cell viability, the alamar blue method is widely accepted for its simple operation and. Bioteks visual abstracts are brief, animated presentations that describe the workflow of a single specific application. Analysis of cell viability by the alamarblue assay request pdf. No qc protocol is recommended for fluorescence since.
The epub format is best viewed in the ibooks reader. Robotic mammosphere assay for highthroughput screening in. It can also be used to study mycobacteria, bacteria, and fungi. Microplate alamar blue assay for staphylococcus epidermidis. Optimized alamarblue assay protocol for drug doseresponse.
Franzblaumicroplate alamar blue assay versus bactec 460 system for highthroughput screening of compounds against mycobacterium tuberculosis and mycobacterium avium antimicrob. Material amount concentration storage stability alamarblue reagent 25ml cat. Pdf optimized alamarblue assay protocol for drug doseresponse. Resazurin microtiter assay plate method for detection of. A simple method to measure cell viability in proliferation. It uses the indicator dye resazurin to measure the. Assessment of cell proliferation with resazurinbased. The celltiterblue cell viability assay is a singleaddition, homogeneous assay that measures cell viability using a redox indicator dye resazurin and allows users to measure changes using fluorescence preferred or absorbance. The protocol with the reagents is not very clear and cites references. Therefore, a lot of research on cell viability assay has been carried out.
Cell proliferation and cell viability analysis in in vitro. Sep 10, 2012 these findings suggest that the alamar blue assay may not be appropriate for studying proliferation in tendonderived cells. In vitro toxicology assay kit, resazurin based tox8. Throughput in tuberculosis drug discovery was extremely limited prior to the introduction of microplatebased susceptibility assays. The cellquantiblue tm assay kit has been specially optimized and formulated to provide a sensitive, convenient and robust assay for cell proliferation and cytotoxicity. Resazurin cell viability assay offers a simple, rapid, reliable, sensitive, safe and costeffective measurement of cell viability. It is a proven safe and nontoxic dye used for quantitative analysis of cell viability and cell proliferation, for. Viable cells retain the ability to reduce resazurin into resorufin. It can be applied in studies concentrating on animal, plant, yeast, and bacteria cells. By monitoring the absorbance at 570 nm and 600 nm, relative metabolic activity for the cells can be determined. The performance of alamar blue and 34,5dimethylthiazol2yl2,5diphenyl tetrazolium bromide mtt cell viability assays in a high throughput format were compared.
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. 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. Alamar blue protocol nov302006 has any one done the cell viability cell toxicity assay using alamar blue. Resazurin, the active ingredient of alamarblue reagent, is a nontoxic, cellpermeable compound that is blue in color and virtually nonfluorescent. Pdf accurate prediction of the adverse effects of test compounds on living systems. Compounds such as tetrazolium salts and alamarblue, which can be reduced by cellular. Calculations assume 100 l final volume per well 96well plate. Specifically, the system incorporates an oxidationreduction redox indicator that both fluoresces and changes color in response to chemical reduction of growth medium resulting from cell growth.
The assay is based on detection of metabolic activity through an oxidationreduction redox indicator, which both fluoresces and changes colour in. Alamar blue, choriocarcinoma, invasion, migration, viability. In vitro toxicology assay kit, resazurin based tox8 bulletin. 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. A novel onestep, highly sensitive fluorometric assay to evaluate cellmediated cytotoxicity. Fluorescence can be read using 544 nm excitation and 590 nm emission wavelengths, or absorbance can be read using a. Alamar blue, cell viability, cytotoxicity, resazurin.
The simple protocol involves adding a single reagent directly to cells cultured in serumsupplemented medium. Comparison of alamar blue and mtt assays for high throughput. Cell proliferation was assessed by comparing the results of an alamar blue assay and two dna quantification assays, cyquant and picogreen, on two human cancer cell lines and two human primary cell sources. Alamarblue assay definition of alamarblue assay by medical. Alamarblue assay for cell proliferation bmg labtech. Plate and culture cells in a clearbottom 96well tissue culture plate. It is a nontoxic, water soluble, redoxsensitive dye that changes from its blue nonfluorescent state to a pinkhighlyfluorescent state upon reduction by viable cells. The absorbance spectrum for reduced and oxidized forms of the resazurin dye are highlighted in figure 1. C protect from light when stored as directed this kit is stable until the expiration date printed on the product. A total of 117 drugs chosen for their wide range of therapeutic areas were screened at 10. Rapid colorimetric assay based on the cleavage of the tetrazolium ring of mtt 34,5dimethylthazolk2yl2,5diphenyl tetrazolium bromide by dehydrogenases in active mitochondria of living cells as an estimate of viable cell number. Compounds were then assessed for the following criteria. Assay principle the celltiterblue assay is based on cellular reduction of. The 100% reduced form of alamarblue was produced by autoclaving controls ie.
Alamarblue cell viability reagent from thermo fisher scientific. Section 3 hazards identification this product is not classified as hazardous and no known health hazard is known to be associated with exposure to this product. The alamar blue assay satisfies most of these criteria once the user. I have a problem understanding connecting absorbance with. Ldh cytotoxicity assay, caspase 37 assay, mttmts assay.
Analysis of cell viability by the alamarblue assay csh protocols. Multiple applications of alamar blue as an indicator of. An external file that holds a picture, illustration, etc. It uses the indicator dye resazurin to measure the metabolic capacity of cellsan indicator of cell viability. The resazurin assay also known as alamar blue assay offers a simple, rapid, and sensitive measurement for the viability of mammalian cells and bacteria. It is a proven safe and nontoxic dye used for quantitative analysis of cell viability and cell proliferation, for cytokine bioassays and in vitro cytotoxicity studies. Just dilute into your cell growth media and place on your cells. Principle of the assay the alphalisa technology allows the detection of molecules of interest in serum, plasma, cell culture supernatants or cell lysates in a very sensitive, quantitative, reproducible, and userfriendly assay. Description the celltiter blue cell viability assay provides a homogeneous. Microplate alamar blue assay maba and low oxygen recovery.