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- Additional protein information
- Analytical tools
Biotinylated Protein Ladder Detection Pack #7727
Biotinylated protein ladder detection packs are designed to detect the molecular weight ladders on Western blots when using the horseradish peroxidase (HRP) based Western detection system. The pack contains biotinylated protein ladders and HRP-linked, anti-biotin antibody. The molecular weight ladders are a mixture of purified proteins covalently coupled to biotin that resolve to 10 bands that have a size range of 9-200 kDa. The anti-biotin antibody is used to detect biotinylated protein ladders on Western blots. The packs have been optimized for chemiluminescent Western detection procedures.
The Anti-biotin, HRP-linked Antibody may cross react with endogenously expressed biotinylated proteins. Careful titration may be needed.
The protein ladder consists of 10 proteins ranging in apparent molecular weight from 9 to 200 kDa. The 9 kDa protein is corresponding to aprotinin purified from bovine lung. The proteins from 20-50 kDa are paramyosin fragments; the higher molecular weight proteins are fusions of maltose binding protein (MBP) with paramyosin or paramyosin/lacZ fragments. The 200 kDa protein is corresponding to alpha-2-macroglobulin purified from human plasma.
The anti-biotin HRP-linked antibody should only be used to detect the marker and not other biotinylated proteins.
Notes on Use
(a) Thaw protein ladder on ice. Mix well and aliquot the desired amount of the protein ladder (10 µl for mini-gels and 20 µl for full length gels) into a separate tube.
(b) Heat the ladder to 95-100ºC for 2 minutes.
(c) After a quick microcentrifuge spin, load directly onto gel. To ensure uniform mobility, load an equal volume of 1X Reducing SDS Loading Buffer into any unused wells.Storage: Biotinylated Protein Ladder: Store at -20°C.Anti-biotin HRP-linked Antibody: Store at -20°C.
- Perform chemiluminescent detection according to the manufacturer’s instructions except, at the stage of secondary antibody incubation with the blot, concurrently add the anti-biotin antibody (provided) at a dilution of 1:1000. Incubate for one hour at room temperature with gentle agitation.
- Proceed as usual with the detection procedure.
For Research Use Only. Not For Use In Diagnostic Procedures. Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.