Application
Western blot
Sample
Hamster Cell lysate - whole cell (chinese hamster ovaries)
Loading amount
20 µg
Specification
chinese hamster ovaries
Treatment
cells were overexpressing a VP16 fusion protein
Gel Running Conditions
Reduced Denaturing (4-12%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
Other product details
Dilution
1/1000
Incubation time
12 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: 3% milk
Secondary antibody
Name
Non-Abcam antibody was used: goat anti rabbit IRdye 700
Host species: Goat
Clonality: Polyclonal
Conjugation: IRDye® 700DX
Host species: Goat
Clonality: Polyclonal
Conjugation: IRDye® 700DX
Dilution
1/30000
Detection
Detection method
IR imaging
Exposure
3 minute(s) and 0 second(s)
Bands
Specific: 67, 55 kDa Non-specific: 60, 50, 32 kDa
Positive control
a lysate with a VP16 fused SRF served as positive ctrl, as this had already been detected with the VP16 AB.
Negative control
CHO cells overexpressing an empty vector
Additional data
Additional Notes
the western blot was transferred to a Nitrocellulose membrane, primary AB incubation overnight at 4°C. For the imaging the infrared imager Odyssey (Licor) was used.
In generell, the blot was clean, with an appropriate negative and positive control, specific bands are easy to distinguish from the 3 unspecific bands.
It shows in the left lane VP16 fused SRF (full length, slighly above the
unspecific band, approximately 68 kDa), 2nd lane a deletional mutant of SRF
fused to VP16 (slightly below the 60 kDa unspecific band), 3rd lane empty,
4th lane empty vector transfected CHO-cells. The "stronger" unspecific band
is at approximately 60 kDa.
In generell, the blot was clean, with an appropriate negative and positive control, specific bands are easy to distinguish from the 3 unspecific bands.
It shows in the left lane VP16 fused SRF (full length, slighly above the
unspecific band, approximately 68 kDa), 2nd lane a deletional mutant of SRF
fused to VP16 (slightly below the 60 kDa unspecific band), 3rd lane empty,
4th lane empty vector transfected CHO-cells. The "stronger" unspecific band
is at approximately 60 kDa.
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提交于 Apr 02 2008