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Prestoblue

Manufactured by Promega
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PrestoBlue is a resazurin-based solution used to assess cell viability and proliferation in cell-based assays. It measures the reducing environment of the living cells, which indicates their metabolic activity. PrestoBlue provides a simple, quantitative method for determining cell number and monitoring cellular responses to various treatments or stimuli.

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Lab products found in correlation

Prestoblue, by Thermo Fisher Scientific (2 mentions) Luciferin substrate, by Promega (1 mentions) Multidrop combi liquid dispenser, by Thermo Fisher Scientific (1 mentions) Prism v7, by GraphPad (1 mentions) Synergy ht multi detection microplate reader, by Agilent Technologies (1 mentions) Ultra low attachment 384 well plates, by Corning (1 mentions) Celltiter glo 3d reagent, by Promega (1 mentions) Caspase glo 3 7 assay, by Promega (1 mentions) Celltiter glo, by Promega (1 mentions) Celltiter glo luminescent cell viability assay, by Promega (1 mentions) Glomax multi microplate reader, by Promega (1 mentions)

7 protocols using prestoblue

1

Quantifying Soluble TGF-β Bioactivity

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Quantification of soluble-TGF-β bioactivity was determined for co-culture conditions using transformed mink lung cells (TMLC) that have been genetically modified to produce luciferase under control of the TGF-β -responsive plasminogen activator inhibitor-1 (PAI-1) promoter [Abe et al., 1994 (link); Wipff et al., 2007 (link)]. TMLC were a kind gift from Dr. Daniel Rifkin in the Department of Cell Biology at the New York University School of Medicine. TMLC (8 × 103/cm2;n ¼ 3) were grown overnight before being exposed to the conditioned medium from co-culture experiments for one day. TMLC cultured in 1% serum medium were utilized as basal controls. Afterwards, cells were assayed for metabolic activity using PrestoBlue then lysed and luciferase activity was assessed by light production from a luciferin substrate (Promega, Madison, WI, USA) using a luminometer (Perkin Elmer).
Ekwueme E.C., Shah J.V., Mohiuddin M., Ghebes C.A., Crispim J.F., Saris D.B., Fernandes H.A, & Freeman J.W. (2015). Cross-Talk Between Human Tenocytes and Bone Marrow Stromal Cells Potentiates Extracellular Matrix Remodeling In Vitro. Journal of cellular biochemistry, 117(3), 684-693.
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2

BaF3 Cell Viability Assay

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The day before treatment, stable construct expressing BaF3 cells (1×104) were seeded in 96-well plates in medium with or without IL3. Cells were treated with DMSO or inhibitors (1 nM to 10 μM) in the presence or absence of IL3 for 72 hr. Cell viability was determined using PrestoBlue (Promega, Madison, WI). Two independent experiments, each in duplicate, were performed.
Cheung L.W., Yu S., Zhang D., Li J., Ng P.K., Panupinthu N., Mitra S., Ju Z., Yu Q., Liang H., Hawke D.H., Lu Y., Broaddus R.R, & Mills G.B. (2014). Naturally occurring neomorphic PIK3R1 mutations activate the MAPK pathway dictating therapeutic response to MAPK pathway inhibitors. Cancer cell, 26(4), 479-494.
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3

Screening Patient-Derived Spheroids Against Oncology Drugs

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Low-passage patient-derived cells, cultured in serum-free conditions, were plated using a Multidrop Combi liquid dispenser (ThermoFisher Scientific) into 384-well ultra-low attachment plates (Corning) and cultured for 24 hours to allow for robust spheroid formation. Approved Oncology Drugs Set VIII compounds were added to wells either as a single shot dose of 10 µM or in a 6-point dose response in duplicate using the Biomek FX liquid handling platform. Each experiment was carried out twice, with vehicle (5% DMSO) and cell death controls (25 µM benzethonium chloride).
After 96-hour incubation in a Cytomat 6000 carousel incubator, PrestoBlue or CellTiter-Glo 3D reagent (Promega) was added to wells at volumes recommended by the manufacturers using a Multidrop Combi. Cell viability was measured using a Synergy HT Multi-Detection Microplate Reader (BioTek) and dose-response curves and IC50 values calculated using GraphPad Prism v7.0.
Taylor J.T., Ellison S., Pandele A., Wood S., Nathan E., Forte G., Parker H., Zindy E., Elvin M., Dickson A., Williams K.J., Karabatsou K., McCabe M., McBain C, & Bigger B.W. (2020). Actinomycin D downregulates Sox2 and improves survival in preclinical models of recurrent glioblastoma. Neuro-Oncology, 22(9), 1289-1301.
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4

RNA Editing Impacts Drug Sensitivity

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The IL-3–dependent Ba/F3 parental cell line was maintained in RPMI 1640 medium containing 5% FBS and 5 ng/ml of IL-3. The spontaneously transformed Ba/F3 cell line was maintained in RPMI 1640 medium containing 5% FBS without IL-3. Stable Ba/F3 cell lines expressing the wild-type and edited genes were obtained and maintained by selection of puromycin (0.6 ug/ml) and IL-3 withdrawal. The 145-compound library was purchased from the John S. Dunn Gulf Coast Consortium for Chemical Genomics (Houston, TX). These compounds were dissolved in DMSO as 10 mM stock solutions. The day before treatment, cells (1×104) were seeded in 96-well plates in medium with or without IL-3. Eight serial dilutions of each compound were prepared in media, and final drug concentrations ranged from 0 to 10 μM. Cells were treated with DMSO or drug compounds in the presence or absence of IL-3 for 72 hr. Cell viability was determined using PrestoBlue (Promega, Madison, WI) for mitochondrial dehydrogenase activity. Drug screening was repeated independently to ensure the reproducibility of the results.
To comprehensively assess the effects of RNA editing sites on drug sensitivity, we downloaded the drug screening data from CCLE (http://www.broadinstitute.org/ccle/home), and calculated the correlations between the RNA editing level and IC50.
Han L., Diao L., Yu S., Xu X., Li J., Zhang R., Yang Y., Werner H.M., Eterovic A.K., Yuan Y., Li J., Nair N., Minelli R., Tsang Y.H., Cheung L.W., Jeong K.J., Roszik J., Ju Z., Woodman S.E., Lu Y., Scott K.L., Li J.B., Mills G.B, & Liang H. (2015). The Genomic Landscape and Clinical Relevance of A-to-I RNA Editing in Human Cancers. Cancer cell, 28(4), 515-528.
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5

Evaluating MSC Proliferation with PrestoBlue

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Murphy Roths Large MSC proliferation rate was assessed using the PrestoBlue assay (Promega, Charbonnières-les-Bains, France) and following the manufacturer’s recommendations. Briefly, MSCs were seeded at the density of 3,500 cells/cm2 in a 6-well plate 48 h after transfection, in a proliferative medium containing DMEM supplemented with 10% of fetal calf serum, 100 U/ml of penicillin/streptomycin, and 2 mmol/ml of glutamine. After 3 days of culture, MRL MSCs were collected, and the number of viable cells was quantified.
Tejedor G., Contreras-Lopez R., Barthelaix A., Ruiz M., Noël D., De Ceuninck F., Pastoureau P., Luz-Crawford P., Jorgensen C, & Djouad F. (2021). Pyrroline-5-Carboxylate Reductase 1 Directs the Cartilage Protective and Regenerative Potential of Murphy Roths Large Mouse Mesenchymal Stem Cells. Frontiers in Cell and Developmental Biology, 9, 604756.
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6

Determining Cell Viability and Caspase Activity

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Viability was measured at 4 and 24 h after nsPEF treatment using the resazurin-based metabolic assay Presto Blue (Life Technologies, Grand Island, NY, USA). Plates were read with a Synergy 2 microplate reader, with excitation/emission settings at 530/590 nm.
In certain experiments, concurrently with cell viability, caspase-3/7 activity was measured. We first recorded fluorescence (Presto Blue/viability) and, then, added the Caspase-Glo 3/7 assay (Promega Corporation, Madison, WI, USA) according to manufacturer’s instructions.
Samples were measured in triplicates, the data were averaged, corrected for the background, and considered as a single experiment.
Rossi A., N. Pakhomova O., Mollica P.A., Casciola M., Mangalanathan U., G. Pakhomov A, & Muratori C. (2019). Nanosecond Pulsed Electric Fields Induce Endoplasmic Reticulum Stress Accompanied by Immunogenic Cell Death in Murine Models of Lymphoma and Colorectal Cancer. Cancers, 11(12), 2034.
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7

Parallel Metabolic and ATP Assays

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Cell viability was measured using PrestoBlue® to assess mitochondrial function and Promega CellTiter-Glo® to assess total ATP production. Neither viability assay in isolation provides a reliable indication of cell health, but when measured in parallel can be used to monitor metabolic activity and energy production. Following the 24 h treatment with APAP or conditioned media, 10% (v/v) PrestoBlue® (A-13262; Life Technologies Paisley, Renfrew, UK) was added to cell culture medium and incubated for 30 min, before measuring the fluorescence signal on a GloMax Multi-Microplate Reader (Promega, Southampton, UK). Total cellular ATP levels were determined on the same cells using the CellTiter-Glo® Luminescent Cell Viability Assay (G7570; Promega, Southampton, UK) and read on the GloMax Multi-Microplate Reader.
Morgan K., Morley S.D., Raja A.K., Vandeputte M., Samuel K., Waterfall M., Homer N.Z., Hayes P.C., Fallowfield J.A, & Plevris J.N. (2023). Metabolism of Acetaminophen by Enteric Epithelial Cells Mitigates Hepatocellular Toxicity In Vitro. Journal of Clinical Medicine, 12(12), 3995.
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