Application
Western blot
Loading amount
20 µg
Gel Running Conditions
Reduced Denaturing (Precast Gel 4-12%, Criterion TGX (Biorad))
Sample
Mouse Tissue lysate - whole (Brain)
Specification
Brain
Treatment
Tissue was mixed in Rippa buffer and denaturated in Laemmli buffer 5min ß 95¯C
Blocking step
Milk as blocking agent for 1 minute(s) · Concentration: 0.5% · Temperature: 4°C
Other product details
Incubation time
30 minute(s) · Temperature: 20°C · Diluent: TBST-Milk 0.5%
Dilution
1/2500
Secondary antibody
Name
Non-Abcam antibody was used: Jackson Immunoresearch, anti-Rabbit
Host species: Donkey
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Host species: Donkey
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Dilution
1/10000
Detection
Detection method
ECL+
Bands
Specific: 100 kDa
Exposure
10 minute(s) and 0 second(s)
Negative control
Samples 1,2,3, 5 and 6
Respectively : Liver, spleen, kidney, Lung, Heart from Mouse
Positive control
Sample 4 (Brain)
Additional data
Additional Notes
For protein detection, a Millipore SNAP i.d.™ 2.0 Protein Detection System was used.
Blocking step was realized in 20sec by Actively-driven 30ml of TBST-milk 0.5% through the membrane then anti-TrkA antibody was added (1/2500, 30min, RT) in TBST-milk 0.5%.
Membrane was washed 3 times (20sec) with 30ml of TBST. Secondary antibody was added (1/1000, 10min, RT) in TBST-milk 0.5 then the membrane was washed again 3 times (20sec) with 30ml of TBST.
Detection was completed with ECL+ in a, ImageQuant LAS4000 (GE)
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. Francois Daubeuf
Verified customer
提交于 Mar 05 2015