Application
Western blot
Loading amount
30 µg
Gel Running Conditions
Reduced Denaturing (10%)
Sample
Fruit fly (Drosophila melanogaster) Tissue lysate - whole (0-4hr embryos)
Specification
0-4hr embryos
Treatment
control shRNA and wee shRNA embryos
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 24°C
Other product details
Incubation time
12 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: 5% BSA/TBST
Dilution
1/1000
Secondary antibody
Name
Non-Abcam antibody was used: Amersham Anti-rabbit IgG, Horseradish Peroxidase-l
Host species: Donkey
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Host species: Donkey
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Dilution
1/10000
Detection
Detection method
Pierce ECL
Bands
Specific: 15 kDa
Exposure
1 hour(s), 30 minute(s) and 0 second(s)
Positive control
0-4hr old wee shRNA embryos (lane 2) should display elevated pHH3Ser10 levels relative to 0-4hr old EGFP shRNA embryos (lane 1)
Additional data
Additional Notes
lysis was in 8M urea by homogenization with Bullet blender (Next Advance) PVDF membrane for 1 hour transfer
total HH3 blot below anti-HH3pSer28 blot to monitor loading
I expected stronger signal in lane 2 given observations with other pSer28 antibodies (ab4178 and ab5169) and exact same lysate
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.
DR. Richelle Sopko
Verified customer
提交于 Mar 04 2014