Application
Western blot
Sample
Human Cell lysate - whole cell (HT29 tumour cells)
Loading amount
50 µg
Specification
HT29 tumour cells
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5%
Other product details
Dilution
1/1000
Incubation time
3 hour(s) and 0 minute(s)
Secondary antibody
Name
Non-Abcam antibody was used: Mouse anti Rabbit
Host species: Mouse
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Host species: Mouse
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Detection
Detection method
ECL
Exposure
5 minute(s) and 0 second(s)
Bands
Specific: 65, kDa Non-specific: 55, 80 kDa
Positive control
IVT cdc25A
Additional data
Additional Notes
Secondary antibody used at a 1:10,000 dilution in 5% milk. I have used the primary antibody at 1:500, 1:1K and 1:2K. I have not blocked for longer to get rid of non-specific bands as this was not an issue for me. It should be noted that the antibody does not seem to be very good at detecting different levels of cdc25A in whole cell lysates.
Abcam response
Thank you for your valuable feedback. The description of the results indicates that this antibody works very well for detection, but is not sensitive enough to quantify changes in amount of Cdc25A antigen in western blotting.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.
MR. Paul Eve
Verified customer
投稿 Jan 09 2007