BiobankCell BiologyBio-Analysis

Reactive Oxygen Species

Understanding biology and disease processes is an incredible challenge. Cell-based assays focusing on signalling, function, morphology, to name but a few enable to get further insight in biology and disease. Cell-based assay is here referred to as any assay measuring or detecting effects, elements or parts of a living cell.

Reactive oxygen species (ROS) are chemical species containing oxygen such as peroxides, superoxide, hydroxyl radical, and singlet oxygen. ROS are chemically active.   In a biological context, ROS are formed as a natural byproduct of the normal metabolism of oxygen and have important roles in cell signaling and homeostasis. During (cellular) stress, ROS levels increase dramatically resulting in significant cellular damage known as oxidative stress. Exogenous ROS can be produced from pollutants, tobacco,  drugs, xenobiotics, or radiation.

Intracellular, ROS are produced through multiple mechanisms depending on the cell model, the major sources being NADPH Oxidase (NOX) complexes in cell membranes, mitochondria, peroxisomes, and endoplasmic reticulum. Mitochondria convert energy for the cell into a usable form, adenosine triphosphate (ATP). ATP is produced via oxidative phosphorylation and involves the transport of protons (hydrogen ions) across the inner mitochondrial membrane by means of the electron transport chain. In the electron transport chain, electrons are passed through a series of proteins via oxidation-reduction reactions, with each acceptor protein along the chain having a greater reduction potential than the previous. The last destination for an electron along this chain is an oxygen molecule. Under normal conditions, the majority of oxygen is reduced to produce water. Only small amounts of ROS are produced as a normal product of cellular metabolism. In particular, one major contributor to oxidative damage is hydrogen peroxide (H2O2), which is converted from superoxide that leaks from the mitochondria.  While ROS are thus produced as a product of normal cellular functioning, excessive amounts can cause deleterious effects. 

Effects of ROS on cell metabolism and cellular function are well documented and include a broad diversity of biological functions: ROS are reported to play a role in Apoptosis, in the induction of host defense genes and in the mobilization of ion transport systems. Platelets involved in wound repair and blood homeostasis blood homeostasis release ROS to recruit additional platelets to sites of injury. ROS also play a role within the adaptive immune system via stimulating the recruitment of leukocytes. Furthermore, ROS are reported  in a variety of inflammatory responses related to cardiometabolic disease such as NAFLD-NASH, T2DM and Cardiovascular disease. ROS induces chronic inflammation by the induction of COX-2, inflammatory cytokines, chemokines and pro-inflammatory transcription factors.

In general, harmful effects of reactive oxygen species on the cell are most often:

  • Damage of DNA or RNA
  • Oxidation of polyunsaturated fatty acids in lipids
  • Oxidations of amino acids in proteins
  • Oxidative deactivation of specific enzymes by oxidation of co-factors

For further information on these assays and our standard experimental set-up, please download the CellMade Cytometry Assays – Technical Information and General Instructions leaflet.

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