Total DNA in brain tissue was extracted using a DNA extraction kit (AidLab, China) and quantified with a NanoDrop 2000 instrument (Thermo Scientific, USA). Equivalent amounts of DNA were digested with a DNA digest mix that consisted of benzonase (Sigma-Aldrich, USA), phosphodiesterase I (Sigma-Aldrich, USA) and alkaline phosphatase (Sigma-Aldrich, USA) for 6 h at 37 °C [50 (link)]. Then the 8-hydroxy-2'-deoxyguanosine (8-OHdG) level in genomic DNA was determined using a DNA Damage ELISA Kit (StressMarq Biosciences, Canada) through a competitive enzyme immune assay, according to the manufacturer’s instructions. The absorbance at 450 nm was recorded using a microplate reader (BioTek Instruments Inc, USA).
Dna damage elisa kit
Manufactured by StressMarq Biosciences
Sourced in Canada
The DNA Damage ELISA Kit is a quantitative assay designed to measure the level of DNA damage in biological samples. It utilizes an enzyme-linked immunosorbent assay (ELISA) format to detect and quantify specific DNA lesions. The kit provides the necessary reagents and protocols to perform the analysis.
Automatically generated - may contain errors
Lab products found in correlation
Nanodrop 2000, by Thermo Fisher Scientific (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Microplate reader, by Agilent Technologies (1 mentions)
Phosphodiesterase 1, by Merck Group (1 mentions)
Dna extraction kit, by Aidlab (1 mentions)
Benzonase, by Merck Group (1 mentions)
Alkaline phosphatase, by Merck Group (1 mentions)
Protein carbonyl assay kit, by Abcam (1 mentions)
Quick dna miniprep plus kit, by Zymo Research (1 mentions)
3 protocols using dna damage elisa kit
1
Quantifying Oxidative DNA Damage in Brain Tissue
2
Urinary Biomarkers for Oxidative Stress
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Urinary biomarkers were normalised to creatinine. 8-OHdG was measured using a DNA Damage ELISA Kit (SKT-120, Stress Marq Biosciences), following a 1:20 dilution. 8-isoprostane was measured following a 1:4 dilution using a Urinary 8-Isoprostane ELISA kit (8isoU1, Detroit R&D). Protein was quantified in 1:10 or 1:20 dilutions using the Pierce BCA Protein Assay Kit (23225, Thermofisher Scientific). Protein carbonyls were analysed using a Protein Carbonyl Assay Kit (ab126287, Abcam) protocol with reagents that were ordered separately. Total oxidized protein (nmol/ml) was normalised to the total protein concentration (mg/ml). Any datapoints that fell outside the limits-of-detection were removed from analysis. There was no effect due to gender or age.
3
Quantifying Oxidative DNA Damage in Tissues
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Oxidative DNA damage was evaluated for 8-OhDG damage using a DNA damage ELISA kit (StressMarq Biosciences Inc.) (Fialkowski et al., 2021) . DNA was extracted from packed red blood cells (PRBCs) and frozen tissue samples (liver, gonad) using a commercially available DNA extraction kit (Zymo quick-DNA miniprep plus kit) as described previously. Extracted samples were stored at 4 ℃ until digestion. Samples were digested at a standardized concentration of ~200 ng/ul (PRBCs, gonad) or 400-500 ng/ul (liver) using a modified digest mix protocol by Quinlivan and Gregory (2008) and stored at -20 ℃ until use in DNA damage plate (Quinlivan and Gregory, 2008) . Digested samples were tested for 8-OhDG damage using a DNA damage ELISA kit at 12x dilution for blood and gonad samples and a 10x dilution for liver samples, with all samples run in duplicate following the manufacturer's instructions. Absorbance was measured using a microplate reader (Epoch2T, Biotech Instruments, Winooski, VT, USA). 8-OhDG concentration was standardized relative to total DNA concentration and reported in ng/µL.
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