Log In Sign up
The largest database of trusted experimental protocols

Multitox fluor multiplex cytotoxicity assay

Manufactured by Promega
Visit Supplier
Sourced in United States

The MultiTox-Fluor Multiplex Cytotoxicity Assay is a fluorescence-based assay that measures multiple cytotoxicity endpoints in a single well. The assay utilizes two fluorogenic peptide substrates to measure the activities of a live-cell protease and a dead-cell protease as indicators of cell viability and cytotoxicity, respectively.

Automatically generated - may contain errors

Lab products found in correlation

Cytation 3, by Agilent Technologies (2 mentions) Cytotox 96 non radioactive cytotoxicity assay, by Promega (2 mentions) Dtx880 multi mode microplate reader, by Beckman Coulter (2 mentions) Opti mem media, by Thermo Fisher Scientific (2 mentions) Rpmi 1640 medium, by Lonza (1 mentions) Lb 941, by Berthold Technologies (1 mentions) Glomax multi detection system luminometer, by Promega (1 mentions) 96 well plate, by Corning (1 mentions) Countess autocounter, by Thermo Fisher Scientific (1 mentions) Trypan blue, by Thermo Fisher Scientific (1 mentions) Dmem f12 medium, by Merck Group (1 mentions) Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions) Horse serum, by Merck Group (1 mentions) Waymouth s mb 752 1 medium, by Thermo Fisher Scientific (1 mentions) Dmem medium, by Thermo Fisher Scientific (1 mentions) Propidium iodide, by Cayman Chemical (1 mentions) Opti mem reduced serum medium, by Thermo Fisher Scientific (1 mentions) Bright glo luciferase assay system, by Promega (1 mentions) Hela cell, by American Type Culture Collection (1 mentions) Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

MultiTox-Fluor Multiplex Cytotoxicity Assay Kit (8 citations) MultiTox-Fluor Cytotoxicity Assay (5 citations) Cytotoxicity assays (2 citations)

26 protocols using multitox fluor multiplex cytotoxicity assay

1

Assessing ECFC Cytotoxicity Using MultiTox-Fluor

Check if the same lab product or an alternative is used in the 5 most similar protocols
The ratio of dead/live ECFCs was determined using the MultiTox-Fluor Multiplex cytotoxicity assay (Promega, WI). Briefly, ECFCs were seeded in a 96-well plate at a density of 1×103 cells/well. After 4 h, ECFCs were treated with various concentrations of 1 in triplicates for 72 h prior to the addition of assay substrates directly to the media for 2 h to allow the reaction to complete. The fluorescence was then measured on SpectraMax plate reader at λ400 nm/505 nm and 485 nm/520 nm, to detect the products of live and dead cell-associated proteases, respectively.
Mohyeldin M.M., Akl M.R., Siddique A.B., Hassan H.M, & El Sayed K.A. (2016). The marine-derived pachycladin diterpenoids as novel inhibitors of wild-type and mutant EGFR. Biochemical pharmacology, 126, 51-68.
+ Open protocol
+ Expand
2

Cytotoxicity Assay for Kawasaki Disease

Check if the same lab product or an alternative is used in the 5 most similar protocols
The cytotoxicity assay used was a fluorescence assay measuring the activity of dead-cell protease released from cells with impaired membrane integrity, which is associated with cell death. The cytotoxicity assay was performed according to the manufacturer’s instructions by using a MultiTox-Fluor Multiplex Cytotoxicity Assay (Promega, USA). Briefly, HUVECs were cultured at a density of 0.5 × 104 cells per well in a 96-well plate in growth medium. Cells were incubated with 7.5% serum from KD patients or patients with bacterial infections in RPMI 1640 medium (Lonza, Tokyo, Japan) for a further 24 h in 96-well plates at 37 °C under a 5% CO2 atmosphere. HUVECs stimulated with sera from KD patients were treated with IG and/or PSL and incubated at 37 °C under a 5% CO2 atmosphere for a further 24 h with a final volume of 100 μl per well. Cells were incubated for 1 h with 100 μl per well of 2X MultiTox-Fluor Multiplex Cytotoxicity Assay reagent at 37 °C. The resultant fluorescence from dead cells was measured using a Tristar multimode microplate reader LB 941 (Berthold Technologies) at, 485 nm Ex/520 nm Em. The fluorescence produced was directly proportional to the number of dead cells. Blank values were subtracted, and the increase in activity was calculated on the basis of activity measured from untreated cells. Each sample was measured in duplicate.
Ueno K., Ninomiya Y., Hazeki D., Masuda K., Nomura Y, & Kawano Y. (2017). Disruption of Endothelial Cell Homeostasis Plays a Key Role in the Early Pathogenesis of Coronary Artery Abnormalities in Kawasaki Disease. Scientific Reports, 7, 43719.
+ Open protocol
+ Expand
3

Multiplex Cytotoxicity Assay Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols
Viability and cytotoxicity were measured with a MultiTox‐Fluor Multiplex Cytotoxicity Assay (Promega), according to the manufacturer's instructions. The MultiTox‐Fluor Assay simultaneously measures two protease activities: One is a marker of cell viability using a fluorogenic, cell‐permeant peptide substrate (glycyl‐phenylalanylamino fluorocoumarin; GF‐AFC), and the other is a marker of cytotoxicity (bisalanyl‐alanyl‐phenylalanyl‐rhodamine 110; bis‐AAF‐R110). The live‐ and dead‐cell proteases produce different products, AFC and R110, which have different excitation and emission spectra, allowing them to be detected simultaneously. Results are provided in relative fluorescence units (RFU) measured using a GloMax‐Multi Detection System: Luminometer (Promega, USA). Data are representative of at least three independent experiments done in triplicate.
Harhouri K., Navarro C., Depetris D., Mattei M., Nissan X., Cau P., De Sandre‐Giovannoli A, & Lévy N. (2017). MG132‐induced progerin clearance is mediated by autophagy activation and splicing regulation. EMBO Molecular Medicine, 9(9), 1294-1313.
+ Open protocol
+ Expand
4

Multiplex cytotoxicity assay for ASB-14 on hAEC

Check if the same lab product or an alternative is used in the 5 most similar protocols
MultiTox-Fluor multiplex cytotoxicity assay (Promega, Madison, WI) was used to determine the toxicity of ASB-14 to hAEC. Cells were seeded on tissue culture treated 96-well plate (Corning, Corning, NY) at 167 cell/mm2 concentration. Following overnight culture, cells were incubated with fresh EGM-2F media for 1 h (to remove built up metabolites). Cells were then incubated with ASB-14 (0.012%, 0.0070%, 0.005%, 0.035%, 0.002% w/v) dissolved in 100 µl of culture media for 1 h at 37 °C, 5% CO2. Following incubation with spiked media, cells were incubated with 100 µl of MultiTox-Fluorfor Reagent for 1.5 h at 37 °C, 5% CO2. Fluorescence intensity was measured using Cytation3 (BioTek, Winooski, VT) plate reader, with live (400Ex/505Em nm) and dead (485Ex/520Em nm) cell fluorescence measured. Fluorescence ratio was calculated as relative fluorescence units (RFU) of live cells/RFU dead cells in the same well. Percentage of viable cells was calculated as fluorescence ratio of the sample/fluorescence ratio of 100% live cells (untreated cells). 60 µg/mL digitonin was used as 100% dead control. All data points were generated from the mean of technical triplicates, with N = 6 experimental replicates for each detergent concentration.
Shklover J., McMasters J., Alfonso-Garcia A., Higuita M.L., Panitch A., Marcu L, & Griffiths L. (2019). Bovine pericardial extracellular matrix niche modulates human aortic endothelial cell phenotype and function. Scientific Reports, 9, 16688.
+ Open protocol
+ Expand
5

Evaluating Cisplatin-Induced Cytotoxicity

Check if the same lab product or an alternative is used in the 5 most similar protocols
Mouse primary tubular epithelial cells from wild-type, mutant mice or NRK52E cells were plated in 96-well plate. Transfection of NRK52E cells with siRNA was carried out as described above. After 24 h of transfection, cisplatin was added for 24 h. Cells were harvested and stained with trypan blue (Thermo Fisher #T10282) to visualize dead cells. Cell counts were analyzed in Countess Auto Counter (Invitrogen, C10227). LDH release was quantified using CytoTox 96 Non-Radioactive Cytotoxicity Assay (Promega #G1780). Medium was collected in triplicates, spun down, and incubated in a 96-well plate with the CytoTox reagent for 20–30 min. After adding the stop solution, the absorbance signal was measured at 490 nm in a plate reader. The ratio between dead cell and live cell was calculated through the measurement using MultiTox-Fluor Multiplex Cytotoxicity Assay (Promega #9201). Cells were incubated in assay buffer containing live cell substrate GF-AFC and dead cell substrate bis-AAF-R110 for 30 min at 37 °C. The plate fluorescence was measured using a plate reader as following: viability: excitation ~400 nm; emission ~505 nm. cytotoxicity: excitation ~485 nm; emission ~520 nm.
Guan Y., Liang X., Ma Z., Hu H., Liu H., Miao Z., Linkermann A., Hellwege J.N., Voight B.F, & Susztak K. (2021). A single genetic locus controls both expression of DPEP1/CHMP1A and kidney disease development via ferroptosis. Nature Communications, 12, 5078.
+ Open protocol
+ Expand
6

Polyphenol Inhibition of SEA Activity

Check if the same lab product or an alternative is used in the 5 most similar protocols
The inhibitory effects of polyphenols on SEA activity, using the assay for an enzymatic cleavage, was estimated with the MultiTox-Fluor Multiplex Cytotoxicity Assay (Promega Co., Madison, WI, USA) according to the manufacturer’s instructions [12 (link)]. Spleen cells were plated in individual wells in 96-well plates (2.5 X 105/mL) in a Russ-10 medium and treated with SEA (200 ng/mL) and polyphenol samples (1–25 μg/mL), followed by incubation at 37°C in a 5% CO2 incubator. After incubation for 48 h, the fluorescent emission intensity produced after the cleavage reaction of the glycyl-phenylalanyl-aminofluorocoumarin (GF-AFC) from viable cells (excitation at 355 nm and emission at 523 nm) was determined.
Shimamura Y., Aoki N., Sugiyama Y., Tanaka T., Murata M, & Masuda S. (2016). Plant-Derived Polyphenols Interact with Staphylococcal Enterotoxin A and Inhibit Toxin Activity. PLoS ONE, 11(6), e0157082.
+ Open protocol
+ Expand
7

Cytotoxicity Assay for H9C2 Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
The survival of H9C2 cells were assayed using Multitox-Fluor Multiplex Cytotoxicity Assay (Promega). Briefly, H9C2 cells were plated in 96-well plate under conditions described in “H9C2 cells” section and after the incubation (24 h without viral constructs and 48 h with viral constructs) cells were washed out and DMEM containing 10% FCS and 10 mM 2–4 dinitrophenole (DNP) was added to each well. To measure cell survival 6 h later, the peptide substrate (GF-AFC) that can be cleaved only by live cells was added to the each well. Following 30 min-long incubation at 37 °C, plates were measured using 1420 Multibabel Counter (Victor) plate reader, with excitation at 370 nm and emissions of 480 nm. The percentage of live cells was calculated based on the intensity of fluorescence according to the manufacturer instructions.
Du Q., Jovanović S., Sukhodub A., Ngoi Y.S., Lal A., Zheleva M, & Jovanović A. (2018). Insulin down-regulates cardioprotective SUR2A in the heart-derived H9c2 cells: A possible explanation for some adverse effects of insulin therapy. Biochemistry and Biophysics Reports, 16, 12-18.
+ Open protocol
+ Expand
8

Culturing and Assessing Mouse Leydig Cell Lines

Check if the same lab product or an alternative is used in the 5 most similar protocols
MA-10 mouse Leydig cells (a gift from Dr. Mario Ascoli, University of Iowa, Iowa City, IA) were cultured in Waymouth’s MB752/1 Medium (Life Technologies) supplemented with 15% horse serum (Sigma-Aldrich) and 20 mM HEPES. TM3 mouse Leydig cells (ATCC, Manassas, VA; CRL-1714) were cultured in DMEM/F12 Medium (Sigma-Aldrich) supplemented with 2.5% fetal bovine serum (FBS, Sigma-Aldrich), 5% horse serum, 15 mM HEPES. MLTC-1 mouse Leydig cells (ATCC, CRL-2065) were cultured in RPMI 1640 Medium (Life Technologies) supplemented with 10% FBS. COS-7 African green monkey kidney fibroblast-like cells (ATCC, CRL-1651) were cultured in DMEM Medium (Life Technologies) supplemented with 10% FBS. Cell number was assessed using MultiTox-Fluor Multiplex Cytotoxicity Assay (Promega, G9200), according to the manufacturer’s instructions.
Bu P., Yagi S., Shiota K., Alam S.M., Vivian J.L., Wolfe M.W., Rumi M.A., Chakraborty D., Kubota K., Dhakal P, & Soares M.J. (2017). Origin of a rapidly evolving homeostatic control system programming testis function. The Journal of endocrinology, 234(2), 217-232.
+ Open protocol
+ Expand
9

Antibody-mediated Neurotoxicity in Nodding Syndrome

Check if the same lab product or an alternative is used in the 5 most similar protocols
Human primary neurons (iCell; Cellular Dynamics International) were cultured in laminin-coated (Sigma-Aldrich) 96-well plates at 37°C [5% (v/v) CO2] and in iCell Complete Maintenance Medium (Cellular Dynamics International) at 80,000 cells per well. Before treatment, medium was replaced with Opti-MEM reduced-serum medium (Life Technologies). Neurons were treated with Opti-MEM only (negative control), rabbit IgG polyclonal anti–leiomodin-1 antibody (20 μg/ml), normal rabbit serum (20 μg/ml), patient sera (n = 1, selected at random from patients with nodding syndrome; dilution of 1:200) or the same patient sera sample with all antibodies depleted (n = 1; dilution of 1:200), patient sera (n = 4, selected at random from patients with nodding syndrome with detectable leiomodin-1 antibodies; dilution of 1:200), the same patient samples with leiomodin-1 antibodies depleted (n = 4; dilution of 1:200), or 0.1% (v/v) saponin for 24 hours. Cell death was determined by uptake of propidium iodide (Cayman Chemical) as per the manufacturer’s instructions. Neurons exposed to patient sera with and without antibody depletion or to CSF were tested for toxicity and viability using MultiTox-Fluor Multiplex Cytotoxicity Assay as per the manufacturer’s instructions (Promega). Fluorescence was measured using FlexStation 3 Microplate Reader and SoftMax Pro software.
Johnson T.P., Tyagi R., Lee P.R., Lee M.H., Johnson K.R., Kowalak J., Elkahloun A., Medynets M., Hategan A., Kubofcik J., Sejvar J., Ratto J., Bunga S., Makumbi I., Aceng J.R., Nutman T.B., Dowell S.F, & Nath A. (2017). Nodding syndrome may be an autoimmune reaction to the parasitic worm Onchocerca volvulus. Science translational medicine, 9(377), eaaf6953.
+ Open protocol
+ Expand
10

Transfection and Protein Quantification in HeLa Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
HeLa cells (ATCC, Manassas, VA) were cultured in high glucose Dulbecco’s Modified Eagle’s Medium (ThermoFisher) containing 10% fetal bovine serum and 1% penicillin-streptomycin and maintained at 37°C and 5% CO2. For transfection experiments, cells were plated at 20,000 cells/well in a 96-well plate. After 24 hr, the media in each well was replaced with 150 μL of media containing LNPs. After another 24 hr, assays were performed as described below.
For wells containing EPO mRNA-LNPs, 100 μL of supernatant was removed and measured for EPO concentration using an ELISA assay (Human Erythropoietin Quantikine IVD ELISA Kit, R&D Systems, Minneapolis, MD). Live cell number was quantified using the MultiTox-Fluor Multiplex Cytotoxicity Assay (Promega, Madison, WI). For wells containing Luc mRNA-LNPs, the live cell number was first quantified using the MultiTox-Fluor Multiplex Cytotoxicity Assay. Then, luminescence was measured using the Bright-Glo Luciferase Assay System (Promega). All assays were performed according to manufacturer guidelines, and luminescence/fluorescence was measured using a Tecan infinite M200 Pro microplate reader.
Kauffman K.J., Mir F.F., Jhunjhunwala S., Kaczmarek J.C., Hurtado J.E., Yang J.H., Webber M.J., Kowalski P.S., Heartlein M.W., DeRosa F, & Anderson D.G. (2016). Efficacy and Immunogenicity of Unmodified and Pseudouridine-Modified mRNA Delivered Systemically with Lipid Nanoparticles in Vivo. Biomaterials, 109, 78-87.
+ Open protocol
+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!