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Protein ladder in western blot4/4/2024 ![]() ![]() ![]() Do not heat, dilute, add reducing agent before loading.Īpproximately 0.2~0.4 mg/ml of each protein in buffer (20 mM Trisphosphate pH 7.5 at 25☌), 2% SDS, 1 mM 2-Mercaptoethanol, 3.6 M Urea, and 15% (v/v) glycerol).ĥ μl of 10-180kDa Prestained Protein Ladder resolves 10 bands in 4-20% SDS-PAGE (Tris-glycine buffer) and after Western blotting to PVDF membrane. Western blotting (WB) is a fundamental technique that has Comparable MW detection with competitors’ protein ladders helped scientists understand important signaling pathways, protein associations, protein expression in various cell lines and tissues, and effects of drugs or treatments on protein levels. The ladder is supplied in gel loading buffer and is ready to use. They are used to help monitor electrophoretic separation as well as estimate the size and concentration of the proteins separated in a gel. Protein standards are mixtures of well-characterized natural or recombinant proteins that are loaded alongside protein samples in a gel. The 10-180kDa Prestained Protein Ladder is designed for monitoring protein separated during SDS-polyacrylamide gel electrophoresis, verification of Western transfer efficiency on membranes(PVDF, nylon, or nitrocellulose) and for approximate sizing of proteins. Introduction to Western Blot Protein Standards. Proteins are covalently coupled with a blue chromophore except for two reference bands (one green and one red band at 25 kDa and 75 kDa respectively) when separated on SDS-PAGE (Tris-glycine buffer). The 10-180kDa Prestained Protein Ladder is a three-color protein standard with 10 pre-stained proteins covering a wide range molecular weights for 10 to 180 kDa. Apply more for thicker (>1.5 mm) or larger gel.2~3 μl per well for general Western transfering. Depending on the primary antibody, is sometimes possible to detect also the protein ladder, this does not means that your western blot is wrong or that the bands that you see are unspecific.3 μl or 5 μl per loading for clear visualization during electrophoresis on 15 well or 10 well mini-gel respectively. ![]()
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