Affinity purified goat anti-biotin antibody is conjugated to horseradish peroxidase. This product has been optimized to detect biotinylated protein markers.
For western blots, incubate membrane and diluted antibody in 5% w/v nonfat dry milk, 1x TBS, 0.1% Tween(R) 20 at room temperature for one hour prior to detection. Note: Sodium azide is an inhibitor of HRP-binding and should not be included in the dilution buffer.
Recommended Antibody Dilutions:
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 2 mg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
This antibody may cross react with endogenously expressed biotinylated proteins. Careful titration may be needed.
Chemiluminescence systems have emerged as the best all-around method for western blot detection. They eliminate the hazards associated with radioactive materials and toxic chromogenic substrates. The speed and sensitivity of these methods are unequalled by traditional alternatives, and because results are generated on film, it is possible to record and store data permanently. Blots detected with chemiluminescent methods are easily stripped for subsequent reprobing with additional antibodies. HRP-conjugated secondary antibodies are utilized in conjunction with specific chemiluminescent substrates to generate the light signal. HRP conjugates have a very high turnover rate, yielding good sensitivity with short reaction times.
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