FAQ: Oxidative Stress / Damage
Q: I want to test my samples for oxidative stress, but there are so many oxidative stress markers. How do I know which assays to use?
A: Oxidative stress may be tested either directly, by measuring the presence of various reactive oxygen species, or indirectly by measuring the resulting damage on one or more biomolecules (DNA, RNA, proteins or lipids). The indirect method is more common because of the stability of most markers of oxidative damage and the relative instability of many ROS.
Many journal reviewers are requesting confirmation of oxidative stress via testing of multiple oxidative stress markers. This may be accomplished by testing multiple markers on one biomolecule (e.g. different markers of protein damage), or by testing one marker each on different molecules (e.g. protein, lipid and DNA).
Here are the best assays to begin your oxidative stress studies:
You may also use our Oxidative Stress Assay Selection Guide.
Q: How do I know which species may be detected using each of your OxiSelect™ Oxidative Stress Assays?
A: All of our OxiSelect™ assays are species-independent, which means they will work across all species. This is because the assays are specifically measuring the damage marker, not the biomolecule itself. Specific proteins, for example, may vary across different species, but markers of protein damage such as protein carbonyl and 3-nitrotyrosine are always the same.
Q: I purchased your OxiSelect™ Comet Assay Kit. Would I be able to do a trial run of the kit using regular microscopy slides?
A: No, you cannot use a regular microscopy slide for the Comet Assay. Our Comet slides have been pretreated chemically to increase adhesion. Your cell/agarose droplet will not be able to stay on a regular slide.
Q: Does your Superoxide Dismutase Activity Assay detect all forms of superoxide dismutase?
A: SOD enzymes are classified into three groups: cytosolic Cu/Zn-SOD, mitochondrial Mn-SOD, and extracellular Ec-SOD. Our OxiSelect™ Superoxide Dismutase Activity Assay Kit detects all forms of SOD.
Q: We are using your 8-OHdG DNA Damage ELISA. Once the DNA of our sample has been extracted, does it have to be immediately used in the assay, or can it be frozen for later testing?
A: 8-OHdG is a very stable modification of DNA; you can definitely store the extracted DNA or digested DNA at -80°C for up to 12 months.
Q: What is the minimum amount of DNA I need to run the OxiSelect™ 8-OHdG ELISA Kit?
A: To reliably detect 8-OHdG in your DNA sample, you need a minimum of 2 µg of isolated/digested DNA per well.
Q: What is considered the normal baseline level of 8-isoprostane (8-iso-Prostaglandin F2α) in humans?
A: The total normal 8-iso PGF2α is about 40-100 pg/mL in human plasma and 500-2000 pg/mL in human urine.
Q: I want to analyze my serum samples for protein oxidation. Should I dilute my serum samples with PBS or Reduced BSA?
A: When using our ELISA kits for Protein Carbonyl or AGE, you must first determine the protein concentration of your serum using a BCA or Bradford assay, and then dilute the sample to 10 ug/mL in PBS, not in reduced BSA.
If you are using our AGE ELISA Kit, further dilution may be needed if the OD of your sample is higher than the highest point on the standard curve. This time mix the 10 µg/mL sample with 10 µg/mL reduced BSA so that all wells are coated at a final protein concentration of 10 µg/mL.
