Related Experiment Videos
Anti-ROS-DNA monoclonal antibody as molecular probe for oxidative DNA damage
1Department of Biochemistry, Faculty of Medicine, Aligarh Muslim University, India.
Abstract:
Modification of 400 bp (approximate size) calf thymus DNA with OH radical resulted in lowered Tm, modification of thymine (58.3%), guanine (20%) and single strand breaks. Monoclonal antibodies (mAb) generated against ROS-DNA were of IgG1 subclass. The mAb showed strong binding to ROS-DNA and ROS-modified bases and polymers, in particular, of thymine. The mAb, therefore, preferentially recognizes ROS-modified epitopes on nucleic acids. Distinct binding to DNA isolated from aged, but not from normal humans by the monoclonal antibody was observed. The antibody effectively recognized oxidative lesions in DNA from cancer patients. These studies demonstrate the potential application of the mAb as an immunochemical probe to detect oxidative DNA lesions.
Insights
Monoclonal antibodies (mAbs) targeting reactive oxygen species (ROS)-modified DNA show promise for detecting oxidative DNA damage. These antibodies specifically bind to ROS-modified nucleic acids, aiding in the identification of lesions in aged and cancer patient DNA.
Area of Science:
- Biochemistry
- Molecular Biology
- Immunology
Background:
- Oxidative stress causes DNA damage, contributing to aging and diseases like cancer.
- Reactive oxygen species (ROS) induce modifications in DNA bases and strand breaks.
- Developing specific biomarkers for oxidative DNA lesions is crucial for diagnostics.
Purpose of the Study:
- To generate and characterize monoclonal antibodies (mAbs) against ROS-modified DNA.
- To evaluate the specificity and binding affinity of these mAbs to various DNA modifications.
- To assess the potential of these mAbs as diagnostic tools for detecting oxidative DNA damage in human samples.
Main Methods:
- Calf thymus DNA was modified using hydroxyl radicals (OH radical) to induce oxidative damage.
- Monoclonal antibodies (IgG1 subclass) were generated against ROS-modified DNA (ROS-DNA).
- Antibody binding specificity was assessed using ROS-DNA, ROS-modified bases, polymers, and DNA from aged and cancer patients.
Main Results:
- Hydroxyl radical modification of DNA resulted in decreased melting temperature (Tm), thymine modification (58.3%), guanine modification (20%), and single-strand breaks.
- The generated mAbs exhibited strong binding to ROS-DNA, ROS-modified bases (especially thymine), and polymers.
- The mAbs specifically recognized oxidative lesions in DNA from aged humans and cancer patients, but not from normal human DNA.
Conclusions:
- Monoclonal antibodies against ROS-modified DNA preferentially recognize specific oxidative epitopes.
- These antibodies demonstrate potential as immunochemical probes for detecting oxidative DNA lesions.
- The findings suggest applications in diagnosing age-related diseases and cancer through DNA damage assessment.