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Replication Protein A (RPA) is composed of RPA70, RPA32 and RPA14 (70, 32 and 14 kDa subunits, respectively). RPA binds directly to single-stranded DNA (ssDNA) where it organizes and protects ssDNA during DNA replication, recombination and repair. The inability to replicate or repair DNA breaks leads to cell cycle arrest and apoptosis.
DNA repair proteins act to maintain genome integrity by recognizing, binding to and repairing damaged DNA. Deficiencies in the activity of these proteins are linked to the development of many pathological diseases, including cancer. Understanding and quantifying DNA repair proteins can help elucidate the molecular mechanisms of DNA damage and repair pathways, and understand the damage specificity of a repair protein.
The DNA Repair Protein ELISA Kit advantage
DNA Repair Protein ELISAs offer an improved method for studying DNA repair protein activity. Current techniques used to study DNA damage and repair include EMSAs and Western blots, which are time consuming, do not allow for high-throughput and provide only semi-quantitative results. In contrast, Active Motif’s DNA Repair Kits are DNA-binding ELISAs that eliminate radioactivity and provide quantitative results in less than five hours.
Each kit includes a 96-well plate in which multiple copies of a specific damaged oligonucleotide have been immobilized. When cellular extract is added, the repair protein of interest binds to the damaged DNA. Each well is then incubated with a primary antibody that is specific for the repair protein being studied. Addition of a secondary HRP-conjugated antibody and developing solution provides an easily quantified colorimetric readout (Figure 1).
Key features
- Non-radioactive, colorimetric method
- Quantitative results in less than 5 hours
- 10X more sensitive than gelshift
- High and low-throughput compatible – 96-stripwell format
- Ability to assay both cells and tissues
