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.
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.
Affinity purified goat anti-biotin antibody is conjugated to horseradish peroxidase. This product has been optimized to detect biotinylated protein markers.
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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