The γ-H2AX immunofluorescence assay was performed using the OxiSelect DNA Double-Strand Break Staining Kit (Cell Biolabs Inc., San Diego, CA, USA) according to the manufacturer's instructions. Briefly, hDPCs were fixed with paraformaldehyde (3.7%) and incubated with blocking buffer on an orbital shaker. The cells were then incubated with an anti-phospho-histone (γ-H2AX) antibody solution, followed by a secondary antibody and a Cy3 conjugate solution. The nuclei were stained with Hoechst 33342 (Sigma-Aldrich). Fluorescent images were obtained using laser scanning confocal microscopy (Keyence Corp., Osaka, Japan).
Oxiselect dna double strand break staining kit
Manufactured by Cell Biolabs
Sourced in United States
The OxiSelect DNA Double-Strand Break Staining Kit is a laboratory equipment designed to detect and quantify DNA double-strand breaks. It provides a fluorescent-based method to visualize and analyze DNA damage in cells.
Automatically generated - may contain errors
Lab products found in correlation
Laser scanning confocal microscopy, by Keyence (2 mentions)
Hoechst 33342, by Merck Group (1 mentions)
Hoechest 33342, by Merck Group (1 mentions)
Glass bottomed cell culture dishes, by NEST Biotechnology (1 mentions)
Ix70 fluorescence microscope, by Olympus (1 mentions)
Biorevo bz 9000, by Keyence (1 mentions)
Ts100 microscope, by Nikon (1 mentions)
Xcite 120 lamp, by Excelitas (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Bz x710 microscope, by Keyence (1 mentions)
7 protocols using oxiselect dna double strand break staining kit
1
γ-H2AX Immunofluorescence Assay
2
Assessing DNA Double-Strand Breaks Using Immunofluorescence
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA-DSBs formation was examined using the OxiSelect DNA Double-Strand Break Staining Kit (Cell Biolabs, San Diego, United States). The cells (5×105 cells/well) were cultivated on glass-bottomed cell culture dishes (Nest Biotechnology, China) at 37 °C for 24 h. Next, cell cultures were subsequently preincubated with either 50 μM BSO or 5 mM OTC for 20 h prior to exposure to DMAE-CB. Then, the cells were treated with 0.01 mM DMAE-CB in the presence or absence of BSO or OTC at 37 °C for 24 h. For immunofluorescent staining, the cells were fixed with 3.7% paraformaldehyde in PBS for 10 min at room temperature, washed with cold PBS (4ºC) once, and incubated with ice-cold 90% methanol for 10 minutes at 4ºC. Then the cells were incubated with blocking buffer for 30 minutes at room temperature on an orbital shaker and washed with cold PBS once. Thereafter, cells were incubated with anti-phospho-Histone (γ-H2AX) Antibody Solution, and incubated for 1 hour. After washed 5 times with Wash Buffer (PBST), the cells were incubated with secondary antibody, Cy3 Conjugate Solution for 1 hour at room temperature. Cells were further incubated with 10 mg/ml Hoechest 33342 (Sigma) for 5 min. Specimens were analyzed by laser scanning confocal microscopy (Keyence Co., Osaka, Japan).
3
Quantifying DNA Damage Repair by γ-H2AX Foci
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After incubation for 24 h of 5 × 104 cells per well, DNA double strand breaks (DSBs) were induced in A498 cells using etoposide as described in our previous report [17 (link)] and observed in the presence or absence of E2. The ability to repair DSBs was determined by staining with an anti-γ-H2AX antibody using an OxiSelect DNA Double Strand Break Staining Kit (Cell Biolabs, San Diego, CA, USA) according to the manufacturer’s instructions with some minor changes. Briefly, cells were allowed to recover by removing the DSB inducer for the indicated time; the exposed DSBs (shown as γ-H2AX foci) appeared as green fluorescence and nuclei were counterstained with 4′,6-diamidino-2-phenylindole. These fluorescent images were all detected using an Olympus IX70 fluorescence microscope (Olympus, Tokyo, Japan).
4
Quantifying DNA Double-Strand Breaks
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Phosphorylation of the histone H2A variant (γ-H2AX) is a marker of DNA double-strand breaks, which is the gravest form of DNA damage [21] (link). We investigated the DNA double-strand breaks induced during t-PDT using the OxiSelect DNA Double-Strand Break Staining Kit (Cell Biolabs, Inc.). Cells (TE-11R) were treated with the indicated talaporfin sodium with or without subsequent irradiation, and were stained with an anti-phospho-histone antibody 24 h after treatment. DNA double-strand breaks labeled with FITC-conjugated secondary antibody were assessed using a fluorescence microscope (BZ-9000 BIOREVO, Keyence Corp., Osaka, Japan).
5
Quantifying DNA Damage in Microglia
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA damage was assessed using the OxiSelect DNA Double Strand Break (DSB) staining kit (Cell BioLabs, San Diego, CA). N9 microglia were plated in black-walled 96-well plates at a density of 10,000 cells/well. The following day the cells were exposed to 0, 50, or 100 mJ/cm2∙min for 6 hours using the 96-LED device with half of the wells for each light dosage receiving etoposide (100 uM; Cell BioLabs) during the last 1 hour to serve as positive controls. Cells were then immediately fixed and stained as per kit instructions. Fluorescent staining was visualized on a Nikon TS-100 microscope equipped with a B-2E/C filter cube (Nikon, Melville, NY) with illumination provided by a 120 W light source (XCite 120 lamp; Lumen Dynamics, Mississauga, ON, Canada). Images were captured using a Nikon DS-L1 camera system; illumination intensity and exposure time were kept constant across all images.
6
Evaluating DNA Double-Strand Breaks in ESCC Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A total of 2 × 104 KYSE30 and KYSE180 cells were seeded into 96‐well plates 24 h prior to transfection with the ASO. After transfection for 48 h, KYSE30 and KYSE180 cells were treated with 15 Gy ionizing radiation (IR). After 3 h, γH2AX immunofluorescent staining was performed according to the manufacturer's instructions provided in the OxiSelect™ DNA Double‐Strand Break (DSB) Staining Kit (CELL BIOLABS, INC, San Diego, CA, USA).
7
Quantifying DNA Double-Strand Breaks
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA double-stranded breaks (DSBs) were assessed using the OxiSelect DNA Double-Strand Break (DSB) Staining Kit (Cell Biolabs, San Diego, CA, USA) according to the manufacturer’s instructions with slight modifications. Briefly, 4 × 104 cells/well were seeded in a flat-bottom 96-well plate and incubated overnight. Cells were then treated with DMSO, etoposide or different concentrations of DSA for 80 min. Cells were subsequently fixed with 3.7% formaldehyde for 15 min at room temperature in the dark, washed with PBS, permeabilized with ice cold 90% methanol for 10 min at 4 °C in the dark and washed with PBS. Finally, cells were incubated with blocking buffer containing 1% bovine serum albumin (Sigma-Aldrich, MI, USA) diluted in PBS for 30 min at room temperature on an orbital shaker, then stained with an anti-γ-H2A.X antibody for 1 h at room temperature on an orbital shaker and stained with a fluorescein isothiocyanate (FITC)-conjugated secondary antibody for 1 h at room temperature on an orbital shaker. Fluorescence imaging of γH2A.X was conducted using the Keyence BZ-X710 microscope (Keyence Corporation, Osaka, Japan) at a total magnification of 400×.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!