Then, LDH activity levels were determined using the LDH activity kit. A total of 2 × 103 cells/wells were seeded in a 96‐well plate after 48 h of transfection. LDH activity levels were detected using an ELISA reader (Infinite® M200, Tecan trading AG) at 450 nm.
Infinite m200
The Infinite M200 is a multi-mode microplate reader that provides high-performance absorbance, fluorescence, and luminescence detection. It offers precise and versatile measurement capabilities for a wide range of applications in life science research and drug discovery.
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2 648 protocols using infinite m200
Cell Viability and LDH Assay Protocol
Then, LDH activity levels were determined using the LDH activity kit. A total of 2 × 103 cells/wells were seeded in a 96‐well plate after 48 h of transfection. LDH activity levels were detected using an ELISA reader (Infinite® M200, Tecan trading AG) at 450 nm.
Quantification of NOX and XO Activities
Xanthine oxidase (XO-EC1.17.3.2) activity was estimated by uric acid formation from xanthine [92 (link)]. Cell lysates were added to phosphate buffer (50 mM, room temperature, pH 7.5) supplemented with xanthine (150 µM). Next, the changes in absorbance at 290 nm were measured for one minute immediately after mixing the sample (Microplate reader Infinite M200, TECAN Trading AG, Männedorf, Switzerland). One XO unit was defined as the amount of enzyme required to release 1 µM uric acid per minute. Enzyme activity is shown in microunits per mg of protein.
Enzymatic Assays for Liver Health
Transient Transfection and Luciferase Assay
Evaluating Cytotoxicity of Anti-Cancer Compounds
Cytotoxicity Evaluation of Drugs
Regarding the SRB protocol, after cell seeding and treatment, cells were fixed using an ice-cold 10% trichloroacetic acid solution for 30 min and incubated with a 0.4% SRB solution for 1 h at room temperature. To remove the excess dye, plates were washed three times with tap water and allowed to dry. To quantify protein-bound dye, 200 µL of 10 mM Tris base solution was added to each well, and absorbance readings were performed at 510 nm using an automated microplate reader (Tecan Infinite M200, Tecan Group Ltd, Männedorf, Switzerland). Each experiment was repeated three times, using triplicates.
Cytotoxicity Assays for Cell Lines
Measuring Mitochondrial and Plasma Membrane Potentials
Oxidative Stress Biomarkers in Plasma and Liver
Plasma malondialdhehyde: Plasma concentration of malondialdehyde (MDA) was determined by the thiobarbituric acid assay and spectrophotometrically measured at 535 nm (Infinite M200, Tecan, Switzerland) as previously reported by Manso et al. [16 (link)]. The results were expressed as nmol MDA/ml plasma.
Glutathione Reduced in liver: The levels of reduced glutathione (GSH) were measured by the monochlorobimane fluorimetric assay, previously reported by Kamencic et al., by a fluorometer (390 nm excitation and 510 emission; Infinite M200, Tecan, Switzerland) [17 (link)]. The results were expressed as µmol GSH/g protein.
Superoxide and Hydrogen Peroxide Detection
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