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Xtt cell viability kit

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The XTT Cell Viability Kit is a colorimetric assay used for the quantitative determination of cell viability and proliferation. It measures the metabolic activity of cells by detecting the reduction of the yellow tetrazolium salt XTT to an orange formazan dye.

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XTT kit (2 citations)

33 protocols using xtt cell viability kit

1

Adenovirus-mediated β-catenin Expression Assay

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Adenovirus that expresses mouse wild type β-catenin was purchased from Applied Biological Materials. Cell proliferation was assessed by XTT Cell Viability Kit (Cell Signaling) according to manufacturer’s instructions. LNCaP cells were plated in 96-well plates at 104cells/well. Cells were infected with adenovirus expressing β-catenin (Ad-β-catenin) or null adenovirus (Ad-Null) at multiplicity of infection = 50, maintained in normal culture media with steroid depleted serum and treated with 1nM DHT, 1μM Flutamide, 1μM bicalutamide or 30μM MJC13 for 24 h. After 24 h, XTT solution was added to each well and cells were incubated for 4 h at 37°C, the absorbance was measured using an ELISA reader at a wavelength of 450 nm.
Suh J.H., Chattopadhyay A., Sieglaff D.H., Storer Samaniego C., Cox M.B, & Webb P. (2015). Similarities and Distinctions in Actions of Surface-Directed and Classic Androgen Receptor Antagonists. PLoS ONE, 10(9), e0137103.
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2

Cellular Toxicity Assay for Antiretroviral Nanoparticles

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Cellular toxicity assay was performed on MDM in the presence of EVG native drug, PLGA-EVG NPs, and HS30@PLGA-EVG using an XTT cell viability kit (Cell signaling, Danvers, MA). In this study, MDM were seeded into 96-well plates (2 × 104 in each well). Cells were incubated with 0–20 µM of EVG or equivalent concentrations of PLGA-EVG, and HS30@PLGA-EVG for 24 hours. The cells were washed twice with PBS and replaced with 100 µL fresh media. Subsequently, 50 µL of XTT detection solution, which contains electron coupling solution, was added to each well of the 96-well plate. MDM were incubated with XTT detection solution for 3 hours and absorbance was measured using a plate reader at 450 nm. The percentage of viable cells from treatment groups was calculated by comparing the absorbance with the untreated cells. Data presented are from five replicates.
Gong Y., Chowdhury P., Nagesh P.K., Rahman M.A., Zhi K., Yallapu M.M, & Kumar S. (2020). Novel elvitegravir nanoformulation for drug delivery across the blood-brain barrier to achieve HIV-1 suppression in the CNS macrophages. Scientific Reports, 10, 3835.
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3

Enterococcus Vancomycin Resistance Assay

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Isolates identified as Enterococcus were phenotyped for vancomycin resistance using microbroth dilution according to the CLSI guidelines. ATCC29212 (vancomycin susceptible) and ATCC51299 (vancomycin-resistant) were included in all assays as controls. Isolates were grown to mid-log phase, diluted in culture media to 1×106 CFU/ml, and used to inoculate plates containing vancomycin ranging from 128–2ug/ml. After 24 hours of static growth at 37˚C, optical density was read at 600 nm and MIC was determined by scoring by eye for turbidity.
Vancomycin resistance of biofilms was assayed after establishing biofilms as above. After 24 hours of static growth at 37˚C, planktonic cells were removed by washing three times with sterile water, and then 200 μl of TSBG containing 5 mg/ml, 5 μg/ml, or no vancomycin, and the plates incubated at 37˚C for an additional 24 hours. After washing planktonic cells three times with sterile water, 200 μl sterile water was added to each well and the viability of the cells in the biofilm was assessed using an XTT Cell Viability Kit (Cell Signaling Technology, #9095) according to manufacturer’s protocols, reading absorbance at 450nm 60 minutes after addition of reagents.
Gasparrini A.J., Wang B., Sun X., Kennedy E.A., Hernandez-Leyva A., Ndao I.M., Tarr P.I., Warner B.B, & Dantas G. (2019). Metagenomic signatures of early life hospitalization and antibiotic treatment in the infant gut microbiota and resistome persist long after discharge. Nature microbiology, 4(12), 2285-2297.
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4

Flavonoid-Induced Cell Viability Assay

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Cells (1 X 104) were seeded in 10% FBS containing medium and were allowed to attach to 96-well plates. After 24 h, the medium was replaced with a fresh medium containing 2.5% stripped charcoal serum supplied with either DMSO or flavonoids. The XTT cell viability kit (Cell Signaling Technology, Danvers, MA) was then used and the manufacturer’s protocol was followed to calculate the percentage of cell survival.
Shrestha R., Mohankumar K., Martin G., Hailemariam A., Lee S.O., Jin U.H., Burghardt R, & Safe S. (2021). Flavonoids kaempferol and quercetin are nuclear receptor 4A1 (NR4A1, Nur77) ligands and inhibit rhabdomyosarcoma cell and tumor growth. Journal of Experimental & Clinical Cancer Research : CR, 40, 392.
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5

Microglia Viability and Cytotoxicity Assay

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For viability and cytotoxicity experiments, primary microglia were seeded at a density of 7.5 × 104 cells/well in 150 μL DMEM containing 1% P/S and 1% N-2 supplement (GIBCO) in a 96-well plate and allowed to attach O/N. After pre-simulation with 100 ng/mL lipopolysaccharide (LPS) (InvivoGen) for 3 h, wells were washed once with DMEM and treated with either 1.75 μM ASC, 5 μM Aβ, ASC-Aβ composites (containing 1.75 μM ASC and 5 μM Aβ) or its buffer controls (20 mM Tris (pH 8.0) and 300 mM NaCl (“buffer B”) for ASC and DPBS for Aβ) for 12 and 24 h. Subsequently, LDH release was measured using 50 μL supernatant and a cytotoxicity detection kit (Roche) according to the manufacturer’s protocol. The reaction was stopped with 1 N HCl and absorbance was measured at 490 and 680 nm using a microplate reader (Infinite M200; Tecan).
To determine cell viability, the XTT Cell Viability Kit (Cell Signaling Technology®) was used according to the manufacturer’s protocol. In brief, 150 μL phenol red-free DMEM (GIBCO) per well was mixed with 50 μL of XTT Reagent and 1 μL Electron Coupling Solution and added to the microglia. After 1 h the absorbance was measured at 450 nm with a TECAN microplate reader.
Friker L.L., Scheiblich H., Hochheiser I.V., Brinkschulte R., Riedel D., Latz E., Geyer M, & Heneka M.T. (2020). β-Amyloid Clustering around ASC Fibrils Boosts Its Toxicity in Microglia. Cell reports, 30(11), 3743-3754.e6.
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6

XTT Cell Viability Assay for R55

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Neuronal cell viability upon R55 administration was determined per manufacturer’s instructions using the colorimetric XTT Cell viability Kit (Cell Signaling).
Mecozzi V.J., Berman D.E., Simoes S., Vetanovetz C., Awal M.R., Patel V.M., Schneider R.T., Petsko G.A., Ringe D, & Small S.A. (2014). Pharmacological chaperones stabilize retromer to limit APP processing. Nature chemical biology, 10(6), 443-449.
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7

Measuring LEC Proliferation and Migration

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LECs proliferation assay was performed using the XTT cell viability kit (Cell Signaling Technology). The relative migratory ability of LECs was determined by wound-healing assay. Cells were plated onto 6-well plates and allowed to reach full confluence before a scratch was gently created with a pipette tip. The area devoid of cells was photographed at the time the scratch was created and at 23 hours. The relative distance of LECs migrated into the denuded area was measured.
To assess apoAI uptake by TECs in vitro, we used apoAI labeled with Alexa Fluor 555 and (5/6)-TAMRA-SE, which was exposed to IsoLG and incubated for 4 hours at 37°C. TECs with approximately 70% confluence were starved in serum-free medium overnight then incubated with fluorescence labeled unmodified or IsoLG-modified apoAI ± PPM. After 4 hours, the cell lysates were collected for fluorescent measurements (48 (link)). In complementary studies, TECs grown on slides were costained with actin (Invitrogen) and mounting solution containing DAPI.
Zhong J., Yang H.C., Shelton E.L., Matsusaka T., Clark A.J., Yermalitsky V., Mashhadi Z., May-Zhang L.S., Linton M.F., Fogo A.B., Kirabo A., Davies S.S, & Kon V. (2022). Dicarbonyl-modified lipoproteins contribute to proteinuric kidney injury. JCI Insight, 7(21), e161878.
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8

Cell Viability Assays: XTT and Trypan Blue

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Epithelial cell viability was determined by formazan complex formation using the colorimetric XTT cell viability kit (Cell Signaling, Inc., Danvers, MA) read in situ on a BioTek Synergy2 plate reader. For trypan blue exclusion assays, PBS-washed cells were mobilized with 0.25% trypsin, washed in PBS again, resuspended in 100 µl of PBS, and mixed with equal volumes of 0.4% trypan blue dye. Cell counts in samples were manually determined on a hemacytometer.
Kirkpatrick C.T., Wang Y., Leiva Juarez M.M., Shivshankar P., Pantaleón García J., Plumer A.K., Kulkarni V.V., Ware H.H., Gulraiz F., Chavez Cavasos M.A., Martinez Zayes G., Wali S., Rice A.P., Liu H., Tour J.M., Sikkema W.K., Cruz Solbes A.S., Youker K.A., Tuvim M.J., Dickey B.F, & Evans S.E. (2018). Inducible Lung Epithelial Resistance Requires Multisource Reactive Oxygen Species Generation To Protect against Viral Infections. mBio, 9(3), e00696-18.
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9

T Cell Proliferation Assay

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Purified immune T cells were assessed for their ability to proliferate in response to in vitro restimulation in culture with chlamydial antigen as described previously [16 ] using the XTT Cell Viability Kit according to the manufacturer’s instructions (Cell signaling, Boston, MA). After three days of ex vivo antigen-restimulation, XTT detection solution was added to the T cell mixture and the absorbance read at 450 nm. The stimulation index (SI), the ratio between stimulated and non-stimulated cells, was then calculated.
Pan Q., Pais R., Ohandjo A., He C., He Q., Omosun Y., Igietseme J.U, & Eko F.O. (2015). Comparative evaluation of the protective efficacy of two formulations of a recombinant Chlamydia abortus subunit candidate vaccine in a mouse model. Vaccine, 33(15), 1865-1872.
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10

Cell Proliferation Assay Protocol

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Cell were seeded at 5×103 cells/well in 96-well plates and proliferation determined using a XTT Cell Viability Kit (Cell Signaling Technology) according to the manufacturer’s instructions. Alternatively, cells were seeded on glass cover slips, incubated overnight, fixed and immunostained for the cell proliferation marker, Ki67. Total cell number was evaluated by DAPI staining. The ratio of Ki67 positive cells to the total cell number was determined.
Tang Y., Feinberg T., Keller E.T., Li X.Y, & Weiss S.J. (2016). Snail/Slug-YAP/TAZ Complexes Control Skeletal Stem Cell Self-Renewal and Differentiation. Nature cell biology, 18(9), 917-929.
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