Western blot Abreview for Aromatase peptide

Western blot

Review text: How long did you incubate the protein and antibody mix before adding it to the blot?
First Time: Incubation time of antibody and protein (peptide): two hour at room temperature
Second Time:Incubation time of antibody and protein (peptide): One hour at 37 degree C
How much antibody/protein mix was added to the blot?
Antibody used for the reaction is 1ug and protein (peptide) 5ug (1:5 ratio)
Sample: Mouse Tissue lysate - other (BRAIN)
Loading amount: 15 µg
Specification: BRAIN
Blocking step: Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Other product details
Concentration: 5 µg/ml
Incubation time: 1 hour(s) and 0 minute(s) · Temperature: 30°C

Primary antibody (in addition to 'Aromatase peptide')
Concentration: 1 µg/ml

Secondary antibody
Name: Non-Abcam antibody was used: anti - rabbit
Host species: Goat
Clonality: Polyclonal
Conjugation: Horse Radish Peroxidase
Dilution: 1/20000

Detection method: ECL
Exposure: 2 minute(s) and 0 second(s)
Bands: Specific: 54 kDa Non-specific: 104, 77, 71, 67, 46, 36, kDa

Additional data
Peptide usage 5 times - 1ug of antibody and 5ug of blocking peptide.
Abcam response: The blocking peptide was the same as the immunogen. At the moment we have not found a reason why blocking is not taking place,
We have not had similar complaints for Aromatase peptide (ab51924).
More blocking peptide might be needed in customer's case.

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Submitted Apr 28 2011

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