Related Experiment Video

Updated: Jan 22, 2026

Evaluating In Vitro DNA Damage Using Comet Assay
10:43

Evaluating In Vitro DNA Damage Using Comet Assay

Published on: October 11, 2017

39.4K

DNA Damage Focus Formation Assay

Yoshihiro Fujii1

  • 1Department of Radiological Sciences, Ibaraki Prefectural University of Health Sciences, Ibaraki, Japan. fujiiy@ipu.ac.jp.

Methods in Molecular Biology (Clifton, N.J.)
|July 4, 2019
PubMed

Insights

Advanced techniques detect cellular DNA damage, particularly DNA double-strand breaks (DSBs) from ionizing radiation. The gamma-H2AX and Rad51 focus formation assay visualizes these critical DNA damages.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Radiation Biology

Background:

  • Ionizing radiation induces various DNA damages, with DNA double-strand breaks (DSBs) being the most cytotoxic.
  • DSBs can lead to mutations, chromosomal aberrations, and cell death.
  • Traditional methods for DSB detection include neutral elution, pulse field gel electrophoresis, and premature chromosome condensation.

Purpose of the Study:

  • To describe advanced techniques for visualizing cellular DNA damage, specifically DSBs.
  • To highlight the utility of the gamma-H2AX and Rad51 focus formation assay in DNA damage response studies.

Main Methods:

  • Utilizing cellular immunocytochemistry-based fluorescence detection.
  • Employing the gamma-H2AX focus formation assay.
  • Employing the Rad51 focus formation assay.

Main Results:

  • The described methods allow for the visualization of DSBs within cells.
  • Gamma-H2AX and Rad51 focus formation are indicative of DNA damage response pathways.

Conclusions:

  • Advanced fluorescence-based assays provide powerful tools for studying DSBs.
  • The gamma-H2AX and Rad51 focus formation assays are crucial for understanding cellular responses to DNA damage.
Keywords:
DNA damage focus formation assayDNA double strand breaks (DSBs)Gamma-H2AXImmunocytochemistryRad51

More Related Videos

Assaying DNA Damage in Hippocampal Neurons Using the Comet Assay10:00

Assaying DNA Damage in Hippocampal Neurons Using the Comet Assay

Published on: December 19, 2012

28.1K
A High-Throughput Comet Assay Approach for Assessing Cellular DNA Damage07:57

A High-Throughput Comet Assay Approach for Assessing Cellular DNA Damage

Published on: May 10, 2022

5.9K

Related Concept Videos

DNA Damage Can Stall the Cell Cycle02:36

DNA Damage Can Stall the Cell Cycle

In response to DNA damage, cells can pause the cell cycle to assess and repair the breaks. However, the cell must check the DNA at certain critical stages during the cell cycle. If the cell cycle pauses before DNA replication, the cells will contain twice the amount of DNA. On the other hand, if cells arrest after DNA replication but before mitosis, they will contain four times the normal amount of DNA. With a host of specialized proteins at their disposal,cells must use the right protein at...
3.1K
DNA Damage can Stall the Cell Cycle02:36

DNA Damage can Stall the Cell Cycle

In response to DNA damage, cells can pause the cell cycle to assess and repair the breaks. However, the cell must check the DNA at certain critical stages during the cell cycle. If the cell cycle pauses before DNA replication, the cells will contain twice the amount of DNA. On the other hand, if cells arrest after DNA replication but before mitosis, they will contain four times the normal amount of DNA. With a host of specialized proteins at their disposal,cells must use the right protein at...
10.0K
Overview of DNA Repair02:25

Overview of DNA Repair

In order to be passed through generations, genomic DNA must be undamaged and error-free. However, every day, DNA in a cell undergoes several thousand to a million damaging events by natural causes and external factors. Ionizing radiation such as UV rays, free radicals produced during cellular respiration, and hydrolytic damage from metabolic reactions can alter the structure of DNA. Damages caused include single-base alteration, base dimerization, chain breaks, and cross-linkage.
Chemically...
33.5K
DNA Helicases00:55

DNA Helicases

DNA unwinding helicase enzymes are a type of motor protein. Motor proteins can translocate along filaments or polymers using energy generated from ATP hydrolysis. Helicases are involved in all the important cellular processes where DNA unwinding is required, such as DNA replication, repair, recombination, and transcription. They are present in all living organisms, but vary in their structure, function, and mechanism of action. For example, in prokaryotes, DnaB helicase binds and translocates...
23.9K
Translesion DNA Polymerases02:10

Translesion DNA Polymerases

Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
11.1K
DNA Replication02:40

DNA Replication

DNA replication involves the separation of the two strands of the double helix, with each strand serving as a template from which the new complementary strand is copied.  After replication, each double-stranded DNA includes one parental or “old” strand and one “new” strand. This is known as semiconservative replication. The resulting DNA molecules have the same sequence and are divided equally into the two daughter cells.
Replication in Prokaryotes
DNA replication...
58.9K