Safire2 multi detection microplate reader

Manufactured by Tecan

Sourced in Austria, United States

The Safire2 Multi-detection Microplate Reader is a lab equipment product that measures absorbance, fluorescence, and luminescence in microplates. It provides quantitative analysis of samples in a high-throughput format.

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Lab products found in correlation

Alamarblue cell viability assay kit, by Thermo Fisher Scientific (3 mentions) Z2 coulter counter, by Beckman Coulter (1 mentions) Nadph, by Tecan (1 mentions) Nadph, by Merck Group (1 mentions) H2dcfda, by Thermo Fisher Scientific (1 mentions) Cell counting kit 8 cck, by Beyotime (1 mentions) Dc assay, by Bio-Rad (1 mentions)

Spelling variants of this product

Microplate reader (2786 citations) Safire2 (301 citations) Safire2 microplate reader (97 citations) Multi-Detection Microplate Reader (6 citations)

9 protocols using safire2 multi detection microplate reader

Quantifying Cell Death and Viability

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Cell death was assessed through scoring of detached cells in the culture supernatant after pre-incubation of the cultures with 18α-GA or DMSO for 24 h, 1 h incubation with MMC and a 24 h recovery period in the presence of 18α-GA or DMSO in triplicate using a Coulter Z2 counter (Beckman, Brea, CA, USA). For MMC optimization, cell viability of HFL-1 cell cultures was assessed through scoring of attached MMC-treated cells after 1 h of incubation with various MMC concentrations in triplicate using a Coulter Z2 counter (Beckman, Brea, CA, USA). E8.T4 cells [23 (link)] were continuously supplemented with 1 μg/ml doxocycline (to induce the functional form of Nrf2) or were not supplemented with doxocycline (expression of mutated Nrf2 form) and cell death was assessed in both cultures as described above.
For determination of the survival ratio through crystal violet staining, cells treated as described above were fixed in 4% paraformaldehyde for 20 min and then stained with 0.2% crystal violet in distilled water. Cells were washed with water, air dried and the dye was eluted with 30% acetic acid. Viability was assessed by measuring dye absorbance at 595 nm using the Safire2 Multi-detection Microplate Reader (Tecan, Grödig, Austria).

Lefaki M., Papaevgeniou N., Tur J.A., Vorgias C.E., Sykiotis G.P, & Chondrogianni N. (2019). The dietary triterpenoid 18α–Glycyrrhetinic acid protects from MMC-induced genotoxicity through the ERK/Nrf2 pathway. Redox Biology, 28, 101317.

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Enzymatic NAD(P)H Oxidoreductase Assay

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The recombinant enzymes Fre (derived from E. coli) and XiaP (derived from Streptomyces sp. HKI0576) were tested for NAD(P)H:flavin oxidoreductase activity. Reactions were set up in XiaF assay buffer (20 μM Tris-HCl pH 8.0, 10% glycerol) containing 500 μM NAD(P)H (Sigma-Aldrich), 50 μM FAD/FMN/riboflavin (Sigma-Aldrich) and 1 μM flavin reductase at room temperature, unless stated otherwise. Reaction rates were monitored by following the decrease of NAD(P)H absorbance at 340 nm caused by the oxidation to NAD(P) on the Tecan Safire² multi-detection microplate reader. Assays were carried out in a volume of 100 μl. Reactions without enzyme or flavin served as negative controls. Blanc measurements were performed with XiaF assay buffer und subtracted from each value.

Kugel S., Baunach M., Baer P., Ishida-Ito M., Sundaram S., Xu Z., Groll M, & Hertweck C. (2017). Cryptic indole hydroxylation by a non-canonical terpenoid cyclase parallels bacterial xenobiotic detoxification. Nature Communications, 8, 15804.

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Quantification of Cellular ROS Levels

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Detection of ROS was performed with 2′, 7′-dichlorodihydrofluorescein diacetate H2DCFDA (Molecular Probes, Invitrogen, Carlsbad, CA, USA) as previously described [24 (link)]. HFL-1 cells pre-treated with 18α-GA or DMSO for 24 h and then treated with MMC for 1 h, were resuspended in PBS with or without H2DCFDA at a final concentration of 10 μM (loading buffer) and incubated at 37 °C for 30 min. The loading buffer was then removed; cells were resuspended in pre-warmed complete medium and incubated at 37 °C for 5 min. The absorption and the emission of the oxidation product were measured at 493 and 520 nm, respectively using the Safire2 Multi-detection Microplate Reader (Tecan, Grϕdig, Austria). Each sample was measured in triplicate.

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Cytotoxicity Evaluation via AlamarBlue Assay

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Cell viability was evaluated with an alamarBlue Cell Viability Assay Kit (#88952, Thermo Fisher Scientific Inc. Rockford, IL, USA) according the manufacturer’s instructions. Described briefly, cells were plated in a 96-well plate and exposed to various concentrations of the cytotoxic compounds for an indicated time. The alamarBlue Reagent (10 µl) was added to each well, incubated at 37°C in 5% CO₂ for four hours, and then the plates were measured at 545nm/590nm (Ex/Em) using the Tecan Safire² Multi-detection Microplate Reader (Morrisville NC, USA). Average percentage of inhibition at each concentration was calculated as previously described.

Sun X., Ou Z., Xie M., Kang R., Fan Y., Niu X., Wang H., Cao L, & Tang D. (2015). HSPB1 as a Novel Regulator of Ferroptotic Cancer Cell Death. Oncogene, 34(45), 5617-5625.

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Cell Viability Assay Protocol

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Cell viability was evaluated with an alamarBlue Cell Viability Assay Kit (#88952, Thermo Fisher Scientific Inc. Rockford, IL, USA) according the manufacturer’s instructions. In brief, cells were plated in a 96-well plate and exposed to various concentrations of the cytotoxic compounds for indicated times. The alamarBlue Reagent (10 μl) was added to each well and incubated at 37°C in 5% CO₂ for four hours, and then the plates were measured at 545 nm/590 nm (Ex/Em) using the Tecan Safire² Multi-detection Microplate Reader (Morrisville NC, USA). Average percentage of inhibition at each concentration was calculated as previously described.

Sun X., Ou Z., Chen R., Niu X., Chen D., Kang R, & Tang D. (2015). Activation of the p62-Keap1-NRF2 Pathway Protects against Ferroptosis in Hepatocellular Carcinoma Cells. Hepatology (Baltimore, Md.), 63(1), 173-184.

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Bacterial Membrane Permeability Assay

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For intrinsic permeabilization of bacterial membrane, EtBr uptake was in performed 10 mM potassium phosphate buffer (pH 7.2) as described^{62 (link)} with some modifications. Overnight cultures of P. aeruginosa ATCC 9027 in BHI were transferred to fresh medium and grown to OD₆₀₀ values of 0.5–0.6 (log phase). Cells were harvested and resuspended in 10 mM potassium phosphate buffer (pH 7.2) at a final OD₆₀₀ of 0.12–0.15. The cells (40 µl) were added to 50 μl of 12 µM EtBr, followed by the peptide (20 µg/ml final) samples after 30 s. Excitation and emission wavelengths were set at 545 and 600 nm, respectively. The increase in fluorescence as a result of partitioning of EtBr into the cytosol was measured 20 min after the addition of peptides using a fluorescence spectrophotometer (Tecan Safire² Multi-detection Microplate Reader). Membrane potential was measured by DiSC₃(5) as described.^{63 (link)} Briefly, mid-log phase bacteria were suspended in 5 mM HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) buffer containing 20 mM glucose (OD = 0.05) and DiSC₃(5) (1 µM final) was added for 1 h at room temperature. Then KCl was added at 0.1 M of final concentration. After addition with peptide (20 µg/ml) or TX-100 (final 0.1%), fluorescence was excited at 622 nm (bandwidth 5 nm) and monitored at 670 nm for emission (bandwidth 20 nm).

Chung E.M., Dean S.N., Propst C.N., Bishop B.M, & van Hoek M.L. (2017). Komodo dragon-inspired synthetic peptide DRGN-1 promotes wound-healing of a mixed-biofilm infected wound. NPJ Biofilms and Microbiomes, 3, 9.

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Cytotoxicity Evaluation via AlamarBlue Assay

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Sun X., Ou Z., Xie M., Kang R., Fan Y., Niu X., Wang H., Cao L, & Tang D. (2015). HSPB1 as a Novel Regulator of Ferroptotic Cancer Cell Death. Oncogene, 34(45), 5617-5625.

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CCK8 Assay for Hepatocyte Viability

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Cell viability was evaluated with a CCK8 Cell Counting Kit (#C0042; Beyotime Institute of Biotechnology) according to the manufacturer’s instructions. Briefly, hepatocytes were seeded in a 96-well plate and exposed to tBHP (50 μmol/L) for the indicated times. The 10 μL CCK8 reagents were added to each well and incubated at 37°C in 5% CO₂ for 4 hours, and then the plates were measured at 450 nm using the Tecan Safire2 Multi-detection Microplate Reader (Morrisville, NC).

Zheng Z., Xie J., Ma L., Hao Z., Zhang W, & Li L. (2022). Vitamin D Receptor Activation Targets ROS-Mediated Crosstalk Between Autophagy and Apoptosis in Hepatocytes in Cholestasic Mice. Cellular and Molecular Gastroenterology and Hepatology, 15(4), 887-901.

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Superoxide Dismutase Activity Assay

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Cell pellets were collected and lysed via sonication in RIPA buffer supplemented with proteinase inhibitors. Lysates were centrifuged and the supernatant was collected. Protein concentration in the supernatant was determined with DC assay (Bio-Rad Laboratories, Hercules, USA). The protein concentration was adjusted at 0.4 μg/μL for each sample. 25 μL of each lysate (10 μg) were mixed with either 107.5 μL 16 mM Tris-HCl, pH 8.0, 12.5 μL 300 μM nitroblue tetrazolium chloride (NBT), 12.5 μL 468 μM NADH and 12.5 μL 60 μM phenazine methosulfate (PMS) (Sample) or the same combination without PMS (Sample') and loaded in a transparent 96 well plate. Two wells were also prepared with all components but instead of protein, 25 μL of RIPA buffer were loaded (Blank). The plate was incubated in the dark for 10 min and the absorbance was measured with the Safire² Multi-detection Microplate Reader (Tecan, Grodig, Austria) at 560 nm. To calculate the SOD activity, the following formula was used: [(A0-A1)/A0]*100, where A0 is the absorbance of the Blank and A1 the absorbance of the Sample when the absorbance of Sample' has been subtracted.

Vasilopoulou M.A., Gioran A., Theodoropoulou M., Koutsaviti A., Roussis V., Ioannou E, & Chondrogianni N. (2022). Healthspan improvement and anti-aggregation effects induced by a marine-derived structural proteasome activator. Redox Biology, 56, 102462.

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