DNA damage was measured after harvesting cells, washing and resuspended in 200 μl of PBS, fixing in 2 ml of ice-cold 100% ethanol, and storing overnight at −20°C. Cells were resuspended in 1 ml of blocking buffer (1% FCS/PBS), and incubated with antibody to γ-H2AX (phosphorylated Ser139) (Millipore, USA) in blocking buffer (1:500 dilution) at room temperature for 2 h. Cells were washed, incubated with goat anti-mouse Alex 488 Fab fragment secondary antibody (Invitrogen, New Zealand) (1:400 in blocking buffer for 1 h, at room temperature; dark), washed and resuspended in 1 ml of blocking buffer containing RNase (1 μg/ml) and propidium iodide (PI) (10 μg/ml) for 30 min at room temperature. Cells were analyzed in a Becton Dickinson LSRII and profiles were analyzed with ModFit LT 3 software.
Goat anti mouse alex 488 fab fragment secondary antibody
Manufactured by Thermo Fisher Scientific
Sourced in New Zealand
The Goat anti-mouse Alex 488 Fab fragment secondary antibody is a fluorescently labeled antibody fragment that binds to mouse primary antibodies. It is designed for use in various immunoassay and imaging applications.
Automatically generated - may contain errors
Lab products found in correlation
3 protocols using goat anti mouse alex 488 fab fragment secondary antibody
1
Quantifying DNA Damage via γ-H2AX
2
Measuring DNA Damage Response in Cancer Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells (5 × 105 cells for MCF7 or 5 × 105 cells for H460 per well) were grown in 6-well plates overnight. After 24 h, cells were incubated for 2 h with inhibitors. The concentration used for DJ009 was 5 μM, and the topotecan concentration was 0.5 μM for MCF7 or 1 μM for H460. Cells were harvested, washed and resuspended in 1 mL of blocking buffer (1% FCS/PBS), and incubated with antibody to γ-H2AX (phosphorylated Ser139) (Millipore, USA) in blocking buffer (1:500 dilution) at room temperature for 2 h. Cells were washed, incubated with goat anti-mouse Alex 488 Fab fragment secondary antibody (Invitrogen) (1:400 in blocking buffer for 1 h, at room temperature; dark), washed and resuspended in 1 mL of blocking buffer containing RNase (1 µg/mL) and propidium iodide (PI) (10 µg/mL) for 30 min at room temperature. Cells were analyzed in a Becton Dickinson BD Accuri C6 flow cytometer.
3
Quantification of DNA Damage using γ-H2AX
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
As described in detail previously [17 (link)], cells (106 cells per well) were grown in 6 well plates and incubated with inhibitors for the indicated time. Cells were harvested, washed and resuspended in 1 ml of blocking buffer (1 % FCS/PBS), and incubated with antibody to γ-H2AX (phosphorylated Ser139) (Millipore, USA) in blocking buffer (1:500 dilution) at room temperature for 2 h. Cells were washed, incubated with goat anti-mouse Alex 488 Fab fragment secondary antibody (Invitrogen, New Zealand) (1:400 in blocking buffer for 1 h, at room temperature; dark), washed and resuspended in 1 ml of blocking buffer containing RNase (1 μg/ml) and propidium iodide (PI) (10 μg/ml) for 30 min at room temperature. Cells were analysed in a Becton–Dickinson LSRII and profiles were analysed with ModFit LT 3 software.
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!