Fluorescent microplate reader

Manufactured by Thermo Fisher Scientific

Sourced in United States

The Fluorescent microplate reader is a laboratory instrument designed to measure the fluorescence intensity of samples contained in a microplate. It is used to detect and quantify fluorescent signals, which can be used to analyze a variety of biological and chemical processes.

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Lab products found in correlation

Dual lumi 2 luciferase reporter gene assay kit, by Beyotime (1 mentions) Dcfh da, by Merck Group (1 mentions) Inverted fluorescence microscope, by Olympus (1 mentions) Reactive oxygen species assay kit, by Beyotime (1 mentions) H2dcfda, by Merck Group (1 mentions) Dcfh da, by Beyotime (1 mentions) Mtt dye, by Solarbio (1 mentions)

8 protocols using fluorescent microplate reader

Dual Luciferase Reporter Assay Protocol

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Dual luciferase reporter assay was performed using Dual-Lumi™ II Luciferase Reporter Gene Assay Kit (Beyotime Biotechnology) and a fluorescent microplate reader (Thermo Fisher Scientific). All plasmids were constructed by SyngenTech (Beijing, China) and transfected into cells. Their sequences were listed in
Supplementary Table S2. After 24 h, the cells were lysed, centrifuged (10,000‒15,000
g, 3‒5 min), and the supernatant was retained. A total of 100 μL firefly luciferase detection reagent was added to every 20 μL sample, incubated at room temperature for 5 min and then detected using a fluorescent microplate reader, with the result recorded as fLuc. Subsequently, 100 μL renilla luciferase detection reagent was added to the sample and mixed well. The results were recorded as rLuc using a fluorescent microplate reader. The results were presented as the relative expression levels of fLuc and rLuc (fLuc /rLuc ratios).

Xu S., Cheng Z., Du B., Diao Y., Li Y, & Li X. (2023). LncRNA AP000695.2 promotes glycolysis of lung adenocarcinoma via the miR-335-3p/TEAD1 axis: The role of AP000695.2 in glycolysis of lung adenocarcinoma. Acta Biochimica et Biophysica Sinica, 55(10), 1592-1605.

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Determination of Hydrogen Peroxide Levels

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Determination of H₂O₂ was performed according to the protocol of Beyotime kit (Cat #S0038, Beyotime, China). Briefly, 50 mg of colon tissue fragment was homogenized with 200 ul of lysis buffer and was centrifuged at max speed for 5 min in 4℃. Subsequently, 50 µl of supernatant was mixed with 100 ul test buffer and incubate for 30 min in room temperature. Then A560 was detected by a fluorescent microplate reader (Thermo Fisher, USA). Readings were calculated by the standard curve that was prepared from three series of calibration experiments with five increasing H₂O₂ concentrations (ranging from 1 to 100 µM/l).

Wang Y., Mao Y., Chen X., Huang X., Jiang Z., Yang K., Tian L., Jiang T., Zou Y., Ma X., Xu C., Zhou Z., Wu X., Pan L., Liang H., Zhong L, & Chen C. (2023). Homeostatic control of an iron repressor in a GI tract resident. eLife, 12, e86075.

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Measuring Cellular Oxidative Stress

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The cells were seeded on wells at a density of 1 ×
10⁵ cells/well for 24 h, and 1 mL of DCFH-DA (Sigma-Aldrich,
MO) solution was added following the removal of the culture medium;
it was diluted with serum-free medium at a ratio of 1:1000. Subsequently,
the treated bEnd.3 brain endothelial cells were washed using phosphate-buffered
saline (PBS) buffer to clear the residual DCFH-DA solution following
incubation at 37 °C for 20 min. Lastly, fluorescence was measured
at 488 nm (excitation) and 525 nm (emission) using an inverted fluorescence
microscope (Olympus, Tokyo, Japan) and quantified with a fluorescent
microplate reader (Thermo Fisher, MA).

Hao J., Zhang W., Tong R, & Huang Z. (2021). Febuxostat Prevents the Cytotoxicity of Propofol in Brain Endothelial Cells. ACS Omega, 6(8), 5471-5478.

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Measuring Cellular ROS Levels

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The ROS levels in R2C cells were measured using a reactive oxygen species assay kit (Beyotime Biotech, Shanghai, China). Briefly, cells were placed in a 96-well plate at a density of 4 × 10³ per well and incubated for 24 h. After aspirating the medium and washed by fresh medium twice, a final concentration of 10 μM H2DCFDA (DCFH-DA) was added, and then incubated for 20 min. The solution was immediately removed and the well was washed three times with phenol red-free and serum-free medium, and then the fresh medium was added and the fluorescence intensity was measured with a fluorescent microplate reader (Thermo Fisher Scientific, Waltham, MA, USA) under excitation wavelength at 488 nm and emission wavelength at 525 nm, respectively. In the treated group, cells were subjected to 160 µM of AAPH and 50 µM of different anthocyanins for the standard steps with the incubation time of 5, 10, 40, 80, 160, 320, and 240 min. Each experiment included five replications.

Hu J., Li X., Wu N., Zhu C., Jiang X., Yuan K., Li Y., Sun J, & Bai W. (2023). Anthocyanins Prevent AAPH-Induced Steroidogenesis Disorder in Leydig Cells by Counteracting Oxidative Stress and StAR Abnormal Expression in a Structure-Dependent Manner. Antioxidants, 12(2), 508.

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ROS Detection in Chondrocytes

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ROS detection assays were performed using 2’,7’-dichlorofluorescein-diacetate (H2DCF-DA; Sigma-Aldrich) in chondrocytes cultured with 0, 25, or 50 µg/ml of 7α,25-DHC for 48 h. Fluorescence was measured using a fluorescent microplate reader (excitation 485 nm and emission 520 nm; Thermo Fisher Scientific). Images were acquired by a fluorescence microscope.

Seo J.Y., Kim T.H., Kang K.R., Lim H., Choi M.C., Kim D.K., Chun H.S., Kim H.J., Yu S.K, & Kim J.S. (2023). 7α,25-Dihydroxycholesterol-Induced Oxiapoptophagic Chondrocyte Death via the Modulation of p53-Akt-mTOR Axis in Osteoarthritis Pathogenesis. Molecules and Cells, 46(4), 245-255.

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Intestinal Permeability Measurement

Cited 2 times

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Intestinal permeability was evaluated by determining the amount of FITC‐dextran and D‐lactate in the blood, as previously reported [31 (link), 32 (link)]. Briefly, mice were gavaged with FITC‐dextran (0.4 mg/g body weight) and then euthanized 4 h later. The serum FITC‐dextran concentration was measured using a fluorescent microplate reader with an excitation wavelength of 485 nm and emission wavelength of 535 nm (Thermo Fisher Scientific, Inc.). The serum D‐lactate concentration was detected using a D‐Lactate Colorimetric Assay Kit (cat. no. GMS70095.3, Shanghai Genmed Ltd., Shanghai, China) according to the manufacturer's instructions. The concentration of D‐lactate was calculated using the slope and intercept created from a standard curve.

Yang S., He X., Zhao J., Wang D., Guo S., Gao T., Wang G., Jin C., Yan Z., Wang N., Wang Y., Zhao Y., Xing J, & Huang Q. (2021). Mitochondrial transcription factor A plays opposite roles in the initiation and progression of colitis‐associated cancer. Cancer Communications, 41(8), 695-714.

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Intracellular ROS Detection in Lung Tissue

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The lung tissue was cut into pieces and loaded with the PBS buffer, followed by centrifugation and discarding the supernatant. The tissue suspension was obtained after resuspending using the PBS buffer. DCFH-DA (Cat#S0033M, Beyotime, Beijing, China) was diluted in serum-free medium (dilution ratio 1: 1000), and the final concentration of DCFH-DA was 10 μmol/L. DCFH-DA was added to the tissue suspension, and incubated at room temperature in a dark place for 20 min. The tissue suspension was washed with serum-free medium, and the fluorescence intensity of intracellular ROS was detected by a fluorescent microplate reader (Thermo Fisher Scientific, USA) at 488 and 525 nm of maximum excitation and emission spectra, respectively.

Wu Y., Zhang H., Guan L., Jia X, & Wang M. (2023). S14G-humanin alleviates acute lung injury by inhibiting the activation of NF-κB. Aging (Albany NY), 15(23), 13865-13875.

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ARPE-19 Cell Viability Assay

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The cell viability was detected in this study with MTT assay. MTT dye (Cat No.: M1025, Solarbio, China) was added to the 96-well plate with ARPE-19 cells in different groups and incubated for four hours at 37°C. After replacing the culture media with 150 µL of DMSO to each well, the 96-cell plates were shaken thoroughly until the crystals dissolved. Absorbance was measured at a wavelength of 490 nm using the Thermo Scientific Fluorescent microplate reader (Thermo Fisher Scientific, USA). Three independent experiments were conducted for each group and data were used for advanced analyses.

Wan W., Zhu W., Wu Y., Long Y., Liu H., Wan W., Wan G, & Yu J. (2021). Grape Seed Proanthocyanidin Extract Moderated Retinal Pigment Epithelium Cellular Senescence Through NAMPT/SIRT1/NLRP3 Pathway. Journal of Inflammation Research, 14, 3129-3143.

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